Mitochondrial breakage induced by the herbicide paraquat in cultured human lung cells. Although the intracellular toxic sites of paraquat, a herbicide toxic to human bodies, have remained unclear for a long time, we recently demonstrated paraquat-induced mitochondrial injury in rat lung and liver in vivo. In the present study, cultured human lung cells (A549 adenocarcinoma cell line) which received 0.15 mM paraquat (equivalent to 40 mg/kg i.v.) showed selective mitochondrial breakage at 6-24 hr and died at 36-48 hr. These results suggest that mitochondria are the initial toxic site of paraquat in vitro as well as in vivo in contrast to the previously proposed microsome theory.


Effects of transforming growth factor-alpha on chicken adipocyte precursor cells in vitro. The hyperplastic capacity of adipose tissue resides in a group of fibroblast-like adipocyte precursor cells. There is evidence to suggest that their proliferation and differentiation is regulated by insulin-like growth factor-I (IGF-I) and transforming growth factor-beta (TGF-beta) but there is less information about other growth factors which may also participate in adipocyte precursor cell hyperplasia. Transforming growth factor-alpha (TGF-alpha) is a 50 amino acid polypeptide which has been shown to stimulate proliferation in both neoplastic and normal cell types acting through the epidermal growth factor (EGF) receptor. We have studied the regulation of DNA synthesis and the activity of lipoprotein lipase by TGF-alpha in chicken adipocyte precursor cells in vitro. Both TGF-alpha and EGF stimulated incorporation of [3H]thymidine into DNA in a dose-dependent manner. TGF-alpha was approximately 180-fold more potent than EGF. Addition of TGF-alpha in combination with IGF-I, TGF-beta 1 or platelet-derived growth factor produced a synergistic increase in DNA synthesis. Short-term incubation with TGF-alpha reduced lipoprotein lipase activity by 23%. These results show that TGF-alpha is a potent mitogen in these adipocyte precursor cells and can inhibit their differentiation in vitro and may participate in the regulation of adipose tissue development in vivo.


Effects of the daily administration of gonadotrophin-releasing hormone on the anterior pituitary gland of rats with restricted feeding. To investigate the influence of weight reduction on pituitary function and its modulation by gonadotrophin-releasing hormone (GnRH), female rats were restricted to 10 g food/day for 60 days. GnRH (5 micrograms) or saline (0.2 ml) were administered daily between days 31 and 60 of the period of underfeeding. Underfeeding brought about a decrease in the pituitary gonadotrophin content, serum levels of gonadotrophins and oestradiol, and the number and size of both LH- and FSH-positive pituitary cells. The administration of GnRH to underfed rats produced an increase in the pituitary and serum gonadotrophin levels and the number and size of both LH- and FSH-positive pituitary cells. These observations suggest that underfeeding and/or weight loss diminish the number and activity of the pituitary gonadotrophs, and that daily administration of GnRH both increases the number of gonadotrophs and augments their activity.


Metabolism of dihydrotestosterone to 3 alpha-androstanediol in brain and plasma: effect on behavioural activity in female rats. Six experiments were carried out to determine whether dihydrotestosterone (5 alpha-androstan-17 beta-ol-3-one; DHT) acts to inhibit oestradiol (OE2)-induced lordosis behaviour after its metabolic conversion to 5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-androstanediol, 3 alpha-Adiol). In experiments 1 and 2, ovariectomized rats were treated with several doses of DHT or 3 alpha-Adiol, injected with OE2 and progesterone, and tested for lordosis responsiveness. Significant inhibition of lordosis occurred after a dose of 3 alpha-Adiol which was approximately threefold less than the effective DHT dose. In experiments 3 and 4, plasma concentrations of DHT and 3 alpha-Adiol were measured after the injection of these steroids to ovariectomized rats at doses shown to be both sufficient or insufficient to inhibit lordosis. Behaviourally effective dosages of DHT and 3 alpha-Adiol produced circulating concentrations of 3 alpha-Adiol greater than those produced by behaviourally ineffective doses of DHT or 3 alpha-Adiol. At 30 min after injection of DHT (experiment 3), 78.8% of plasma androgens were in the form of 3 alpha-Adiol, while after injection of 3 alpha-Adiol, only 7.4% were DHT. When plasma DHT and 3 alpha-Adiol were measured at 3, 6, 9 and 12 h after steroid injection (experiment 4), plasma levels of 3 alpha-Adiol produced by the behaviourally subthreshold dose of DHT were significantly lower than levels produced by behaviourally sufficient dosages of DHT or 3 alpha-Adiol. In experiments 5 and 6, concentrations of DHT and 3 alpha-Adiol were measured in five brain regions 1 and 6 h after injection of behaviourally sufficient doses of these steroids to ovariectomized females. At 1 h after injection, similar levels of DHT and 3 alpha-Adiol were measured in DHT- and 3 alpha-Adiol-injected females, and DHT concentrations in the preoptic area were significantly higher in both groups than in any other brain area. At 6 h, animals injected with DHT had significantly higher levels of DHT in all brain areas combined than did 3 alpha-Adiol- or vehicle-injected animals. Brain concentrations of 3 alpha-Adiol were not different between groups injected with DHT, 3 alpha-Adiol or vehicle at this time. In brain, 34.6% of DHT had been converted to 3 alpha-Adiol after 1 h and 53.0% of 3 alpha-Adiol had been converted to DHT.(ABSTRACT TRUNCATED AT 400 WORDS)


Lung to blood passage of human growth hormone after intratracheal instillation: stimulation of growth in hypophysectomized rats. The passage from the lower respiratory tract into the blood of human GH (hGH; M(r) = 22,000) and bovine serum albumin (BSA; M(r) = 67,000) was assessed after intratracheal instillation in adult rats. The plasma level of immunoreactive hGH reached its maximum at 0.5-1 h after instillation and had almost disappeared within 24 h. Higher plasma levels were obtained in male rats than in female rats resulting in a higher total lung passage of hGH in male rats than in female rats (means +/- S.D.; 6.0 +/- 1.7% vs 3.3 +/-1.2%, P<0.01). The plasma level of BSA showed a different pattern, with a maximum at 16-24 h after instillation and a total lung passage of 4.3 +/- 1.7% of the given dose for both sexes. The plasma levels of hGH increased nonlinearly with increasing dose instilled in the dose range 36-720 micrograms/kg body weight. When hGH was instilled daily at a dose of 720 micrograms/kg body weight to hypophysectomized rats for 1 week, they responded with a significant increase in body weight when compared with hypophysectomized control rats (16.8 +/- 4.2 g vs -1.8 +/- 2.4 g, P<0.001). The results demonstrate that, despite their different molecular weights, hGH and BSA pass through the lower respiratory tract into the circulation with similar efficiencies in the rat. However, the lung passage of hGH, unlike that of BSA, showed sexual dependency, an earlier plasma concentration maximum and a tendency of the passage to saturation with increasing dose instilled.(ABSTRACT TRUNCATED AT 250 WORDS)


Cytotoxic factor in the serum of diabetic rats and its increase during pregnancy. Diabetic pregnancy is associated with a high incidence of fetal growth abnormalities which cannot be solely ascribed to fetal hyperglycaemia and hyperinsulinaemia. We therefore examined the possibility of other contributing factors using rats made diabetic with streptozotocin as the experimental model. Blood serum from virgin diabetic rats and, to a much greater extent, from pregnant diabetic rats inhibited [3H]thymidine incorporation by fetal lung cells in culture in a concentration- and time-dependent manner; cell number was also decreased. The cytotoxic activity of the serum was decreased by treatment of pregnant diabetic rats with insulin. Sera from non-diabetic rats and from rats at 6 h and 24 h after the injection of streptozotocin were not cytotoxic. The cytotoxic activity of diabetic rat serum was retained after dialysis and was not destroyed by heating it for 60 min at 60 degrees C. Diabetic rat serum antagonized the stimulatory effects of fetal bovine serum, insulin and insulin-like growth factors on thymidine incorporation by lung cells and inhibited corticosterone production by adrenal cells. Ultrafiltration of diabetic rat serum and high-performance gel permeation chromatography in phosphate buffer suggested that the molecular weight of the cytotoxic factor was approximately 40 kDa. However, gel permeation chromatography in 40% acetonitrile of the cytotoxic eluate from reversed-phase high-performance liquid chromatography using a C18 and C4 column revealed that the cytotoxic factor was a low molecular weight substance, which contained no amino acids. The apparent discrepancy in molecular weights using different separation procedures suggests that the cytotoxic factor is bound to serum proteins. The u.v. spectrum of this factor was different from those of ketone bodies but its exact chemical identity could not be established because of the scarcity of the material. It is suggested that the sera of pregnant diabetic rats and their fetuses contain a cytotoxic factor, which may contribute to fetal developmental abnormalities.


Interaction between thyrotrophin-releasing hormone and the muscarinic cholinergic system in rat brain. TRH increases the pressor response to acetylcholine through an increment in muscarinic receptors. As chronic atropinization produces a similar effect, we hypothesized that both phenomena may be related. The effect of chronic atropine treatment on the TRH content of several brain areas in Wistar rats was studied. Atropine produced significant increases in TRH content in the preoptic and septal areas, while decreases were observed in the hypothalamus and hypophysis. The concentration of TRH in cerebrospinal fluid rose significantly in atropine-treated rats compared with controls. A similar effect was observed with eserine, an acetylcholinesterase inhibitor. Finally, perfusion of brain preoptic area slices from normal rats with Krebs-Ringer solution in the presence of pilocarpine increased basal TRH release significantly and this effect was blocked by atropine. These results are compatible with a muscarinic control on the activity of the central TRH system.


Preparation and characterization of an antibody specific to the rat dopamine D2 receptor. Antibodies specific to the dopamine D2 receptor have been raised in rabbits using synthetic peptides. The resulting antiserum was sensitive to picogram quantities of peptide as measured by enzyme-linked immunoassay and was shown to have a 33% cross-reactivity with partially purified D2 receptor protein. No detectable cross-reactivity with similarly prepared fungal membranes was observed. D2 receptor preparations from normal rat pituitary cells were used in Western blot analysis. Bands of M(r) = 95,000 and 34,000 were detected in these preparations with a third faint band at 120,000. These correspond to the pituitary D2 receptor.


Changes in the plasma concentration of gastric inhibitory polypeptide and other metabolites in response to feeding in sheep. Plasma concentrations of gastric inhibitory polypeptide (GIP)-like activity were determined in sheep before and after refeeding following a 48-h fast. Plasma concentrations increased significantly after feeding, from about 250 pg/ml to about 550 pg/ml. Other metabolites in plasma also increased at this time, reflecting the absorption of nutrients from the gastrointestinal tract. Significant increases were observed in the plasma concentrations of acetate, beta-hydroxybutyrate and triacylglycerol. By comparing the time-courses of the changes in concentration of GIP and other metabolites in plasma, possible sites of secretion and secretagogues of GIP in ruminant animals are proposed. The results demonstrate that GIP is secreted in response to nutrient absorption in adult ruminants and that, as in simple-stomached animals, the absorption of long-chain free fatty acids plays an important role in this secretion.


Influence of lactotroph cell density on prolactin secretion in rats. The relationships between the stimulation of prolactin secretion and proliferation of lactotrophs was studied from a multidisciplinary standpoint in three experimental models. Administration of both oestrogen and sulpiride resulted in a significant increase in prolactin secretion and in the lactotroph population. A single injection of 10 micrograms oestradiol benzoate (OB) induced a twofold increase in the proliferation of lactotrophs (morphometrically as volume density), which increased further (2.5-fold) after three OB injections. Parallel changes were observed in the net counts made on lactotrophs sectioned through the nucleus to avoid possible distortions in volume density caused by hypertrophic cytoplasms. Comparable results were obtained with the mitotic index in the same groups of rats exposed to treatment with colchicine. The effect of sulpiride on proliferation of lactotrophs was also significant (1.7-fold) but less pronounced than in rats treated with oestrogens. The treatments with oestrogen and sulpiride did not stimulate lactotrophic activity in a similar way, as judged by the levels of serum prolactin and the storage patterns of small and big prolactin in pituitary glands. Serum prolactin (mean +/- S.E.M.) in control ovariectomized rats was 4.0 +/- 0.9 micrograms/l and one and three injections of OB raised these levels to 14.4 +/- 5.0 and 28.8 +/- 4.6 micrograms/l respectively. The highest levels of serum prolactin were seen in sulpiride-treated rats (467.2 +/- 28.7 micrograms/l). Striking differences occurred in the pituitary contents of big prolactin, the control values increasing from 5.3 +/- 0.5 to 10.2 +/- 1.3 micrograms/mg after one OB injection and to 14.7 +/- 0.7 micrograms/mg after three OB injections.(ABSTRACT TRUNCATED AT 250 WORDS)


In-vitro DNA synthesis in Leydig and other interstitial cells of the rat testis. Replicative DNA synthesis (125I-labelled iododeoxy-uridine incorporation) was measured in interstitial cells prepared from rat testes and separated by Percoll density gradient centrifugation. Leydig cells were identified by 125I-labelled human chorionic gonadotrophin (hCG) binding and 3 beta-hydroxysteroid dehydrogenase histochemistry. Continuous density gradients indicated that interstitial cell DNA synthesis was not associated with Leydig cells, and was greater in cells from the immature than from the mature rat testis. Fractionation of cells by discontinuous density gradients into Leydig cell-rich and -depleted pools did not result in a similar enrichment of DNA synthesis. Treatment of the adult rat with hCG increased DNA synthesis into both fractions but oestrogen had no effect. DNA synthesis was greater in cells from the immature rat but, in contrast to the adult, in-vivo hCG treatment had no effect, whilst oestrogen decreased synthesis. To characterize the cells synthesizing DNA further, interstitial cells were prepared from testes in which the Leydig cells were depleted by in-vivo treatment with ethane dimethanesulphonate (EDS) or depleted in their germ cells by treatment in utero with busulphan. EDS treatment had no effect on DNA synthesis by the interstitial cells in spite of the 125I-labelled hCG binding being markedly reduced. Similarly, busulphan treatment was also without effects upon DNA synthesis. Fluorescence-activated cell cycle analysis of cells from both fractions from germ cell-depleted testes indicated that only a small proportion (3%) of the interstitial cells were actively dividing and this was almost doubled in cells from the germ cell-depleted immature rat testes.(ABSTRACT TRUNCATED AT 250 WORDS)


Abnormal maternal behaviour in mice previously immunized against progesterone. Anti-progesterone immunization leads to reversible infertility in mice; this can be achieved by passive immunization with a monoclonal antibody to progesterone (DB3), or by active immunization with either a progesterone-protein (bovine serum albumin; BSA) conjugate or anti-idiotype directed against DB3. Recovery of fertility in treated females varied from 39.5 to 75.5 median days after passive or active (progesterone-BSA) immunization respectively. Litter size after the first pregnancy also differed from 8.6 +/- 0.8 to 5.0 +/- 0.6 (mean +/- S.E.M.) per mother after passive or active immunization respectively. When litter size was standardized to a maximum of four pups per litter, aberrant maternal responses were observed in the first 5 days after delivery in 40-70% of the nursing mothers. These responses took the forms of cannibalism and failure to retrieve or to nurse pups and resulted in a high incidence of pup rejection (up to 40%), compared with no rejection in control mothers. When mothers were allowed to keep entire litters, an even higher incidence of pup rejection occurred (51% compared with 8% in controls). There was an apparent relation between the degree of negative maternal behaviour and the progesterone antibody concentration in the circulation during the infertile period. Whereas aberrant behaviour occurred mainly within the first 5 days of lactation, it was significantly reduced thereafter. Aberrant behaviour of the mother towards pups may be a consequence of the presence of residual progesterone antibodies in the circulation which affects the process of progesterone withdrawal at parturition that is essential for the establishment of normal maternal responses to the neonate.


Pituitary-testis function in rats treated neonatally with a gonadotrophin-releasing hormone agonist: short- and long-term effects. Acute and long-term effects of neonatal and prepubertal treatments with an LH-releasing hormone agonist (LHRH-A) were studied in Wistar male rats. Animals injected with D-Ala6-D-Gly10-LHRH ethylamide (2 micrograms/kg per day) or vehicle from days 1 to 15 or from days 16 to 29 were killed at different ages. Treatment between days 1 and 15 induced a decrease in both pituitary FSH and LH content as well as a reduction in plasma FSH and blockade of the response to LHRH. These effects were apparent on day 16 after treatment. Basal and human chorionic gonadotrophin (hCG)-stimulated progesterone and testosterone secretion in vitro was similar in testes from male rats treated with LHRH-A or vehicle. Reduced testicular weight was observed until day 90, whereas puberty, spermatogenesis and fertility were unaffected. The decrease in plasma FSH concentrations after neonatal treatment with LHRH-A was also found in groups of animals killed on day 10 and was possibly the cause of reduced testicular weight, since treatment with FSH from day 1 to day 15 blocked the effect of LHRH-A. Likewise, treatment with LHRH-A from day 1 to day 15 also reduced FSH and LH secretion in males orchidectomized on day 1 of life. Animals injected with LHRH-A from day 15 to day 29 exhibited, at the end of the treatment period, reduced testicular weight, and decreased pituitary gonadotrophin content and plasma FSH concentrations, whereas LH plasma concentrations were normal. In adulthood, the pituitary-testis function did not vary from normal.(ABSTRACT TRUNCATED AT 250 WORDS)


Hypothalamic-pituitary function in neonatally oestrogen-treated male rats. Neonatal oestrogen administration to male rats permanently impaired the function of the pituitary-testicular axis possibly by inhibiting neonatal gonadotrophin secretion. To analyse the hypothalamus and/or pituitary involvement in this inhibition, pituitary responsiveness to acute stimulation with LH-releasing hormone (LHRH) was studied in vivo and in vitro in Wistar male rats injected on day 1 of age with oestradiol benzoate (OB) or olive oil. FSH and LH pituitary content and plasma concentrations were reduced in oestrogenized male rats at days 10 and 16 of age. Likewise, the in-vivo increase in gonadotrophin plasma concentrations after acute stimulation with LHRH was almost completely suppressed in 10- and 16-day-old oestrogenized males. In vitro, the increased secretion of FSH after LHRH stimulation was abolished and the LH response strongly reduced in pituitaries from oestrogenized males. Finally, the effects of neonatal oestrogenization were not abolished by treatment from day 1 to day 15 with an LHRH agonist (0.01 microgram/kg per 12 h). We conclude that in male rats the effects of oestrogenization are due to both a reduction in LHRH endogenous secretion and a decrease in the pituitary responsiveness to LHRH.


Bioactive and immunoreactive FSH and immunoreactive inhibin concentrations in the ovine fetus. The bioactive (B) and immunoreactive (I) pituitary contents/concentrations of FSH, together with the plasma concentrations of B-FSH, I-FSH and I-inhibin were determined in ovine fetuses at days 55, 75, 90 and 135 of gestation (day 145 = term). The pituitary contents and concentrations of B-FSH and I-FSH increased in both sexes with gestational age. The female fetuses had significantly (P < 0.01) higher pituitary contents/concentrations of B-FSH and I-FSH than the male fetuses at days 75 and 135. The pituitary B/I ratios of FSH were not significantly different with age or sex. The plasma concentrations of B-FSH remained relatively constant from days 75 to 135, with no significant differences between sexes or with age. In contrast, the plasma concentrations of I-FSH reached a peak at day 90 and then declined towards term in both sexes. At all gestational ages except day 55, the female fetuses had significantly (P < 0.05) higher plasma concentrations of I-FSH than the males. In both sexes, the plasma B/I ratios of FSH were lowest at day 90 and had increased again by day 135, with the male fetuses having significantly (P < 0.05) higher B/I ratios compared with the female group at days 75 and 135 but not at day 90. At all gestational ages, the plasma concentrations of I-inhibin declined throughout gestation in the female fetuses, whereas in the males they reached a nadir at day 75 and then increased towards term. The concentrations of I-inhibin were significantly (P < 0.01) higher in the male fetuses compared with the females.(ABSTRACT TRUNCATED AT 250 WORDS)


Receptor binding and growth-promoting activity of insulin-like growth factor-I in a bovine mammary epithelial cell line (MAC-T3). Insulin-like growth factor-I (IGF-I) has been known to be mitogenic to a variety of cell types, although a growth-regulatory role for IGF-I on bovine mammary epithelial cells has not been fully investigated. In the present study, we examined the receptor binding of IGF-I and its effect on growth in a bovine mammary epithelial cell line (MAC-T3). Specific receptors for IGF-I were detected on cultured bovine mammary epithelial cells. Competitive binding revealed that half-maximal inhibition of 125I-labelled IGF-I binding by IGF-I was approximately 3 micrograms/l. Dissociation rate constant of the IGF-I receptor was 3.10 +/- 0.06 nmol/l (S.E.M.) with a receptor site concentration of 366 +/- 8 fmol/mg protein for the average of three experiments. IGF-I exerted a positive mitogenic effect on MAC-T3 cells according to both direct DNA assay and thymidine incorporation assay. Moreover, the mitogenic effect of IGF-I on MAC-T3 cells was enhanced by the addition of fetal calf serum in the culture media. The present results suggest that the bovine mammary epithelial cell line (MAC-T3) provides a useful model system with which to study the biological actions of insulin-like growth factors on the bovine mammary secretory tissue in vitro.


Distribution of relaxin between human maternal and fetal circulations and amniotic fluid. Relaxin was measured in maternal blood and amniotic fluid samples at 9-40 weeks and in fetal blood samples at 19-41 weeks of pregnancy. In amniotic fluid, concentrations of relaxin rose from 58 ng/l (geometric mean) at 10 weeks to 142 ng/l at 14 weeks and declined subsequently to 55 ng/l at 22 weeks. In maternal blood, mean relaxin concentrations were ten times greater than in amniotic fluid, and concentrations decreased with gestation. Since there was no significant association between the relaxin concentrations in the two compartments, relaxin in the amniotic fluid may be derived from the decidualized endometrium rather than the maternal circulation, alternatively its metabolism may be different in the two compartments. The absence of detectable concentrations of relaxin in any of the fetal blood samples demonstrates that there is no significant placental transfer or fetal synthesis of this peptide.


Apparent alpha-inhibin subunit immunoactivity in porcine and ovine luteal extracts is due to interference by cytosolic proteases in the assay. Immunoreactive alpha-inhibin (ir-inhibin) was measured in luteal homogenates and subcellular fractions of ovine and porcine corpora lutea (CL) and in pig granulosa cells (GCs), using a sensitive radioimmunoassay specific for the 1-26 amino acid sequence of the N-terminus of the alpha chain of porcine inhibin (p1-26 alpha-inhibin). Inclusion of N-ethylmaleimide (N-EM) and/or EDTA in the immunoassay had no effect on the measurement of p1-26 alpha-inhibin peptide standards, on ir-inhibin levels in ovine follicular fluid and serum, or on ir-inhibin in subcellular fractions of pig GC. Fractionation of porcine GC homogenates on sucrose gradients demonstrated a major particular peak of ir-inhibin (buoyant density, 1.15-1.21 g/cm3) with variable activity in the cytosol. The particulate ir-inhibin peak was released into the cytosol by pretreatment of GC homogenates with the saponin, digitonin, prior to fractionation. Porcine GC extracts contained a protein (M(r) 45,000) which immunoblotted against p1-26 alpha-inhibin antibody. In the absence of inhibitors of proteolysis, apparent ir-inhibin activity was very high in extracts of sheep and pig CL. However, inclusion of N-EM or EDTA in the radioimmunoassay significantly reduced ir-inhibin levels in porcine and ovine CL extracts in a dose-dependent manner. Measurements of peptide tracer integrity indicated that porcine luteal cytosol degraded 125I-labelled p1-26 alpha-inhibin peptide. Subcellular fractionation studies demonstrated high levels of apparent ir-inhibin in luteal cytosol fractions, with only minor activity peaks associated with particulate fractions; however, this material was not releasable by digitonin. Immunoblotting of detergent extracts of porcine luteal particulate fractions failed to demonstrate alpha-inhibin material, and immunocytochemical localization studies of alpha-inhibin in porcine and ovine luteal sections were negative. Our results are consistent with the intracellular packaging/storage of a form of alpha-inhibin (M(r) similar to that of alpha-inhibin subunit precursor) in the porcine granulosa cell. However, luteinization of the porcine follicle was associated with a dramatic fall in ir-inhibin content, and the loss of immunostaining for alpha-inhibin peptides. We conclude that porcine and ovine CL contain little, if any, authentic inhibin. These studies emphasize the importance of excluding proteolytic artefacts when measuring biological peptides in luteal tissue extracts by radioimmunoassay.


Effect of the antiprogestin RU486 on uterine sensitivity to oxytocin in ewes in late pregnancy. The effect of RU486, a synthetic progesterone receptor antagonist, on basal uterine prostaglandin (PG) release and release in response to oxytocin injection has been investigated in late-pregnant sheep (days 135-140 of gestation). Fifteen hours after i.m. injection of RU486 (50 mg; n = 5) or vehicle alone (n = 4), bolus injections of oxytocin (50, 500 and 5000 mU) were administered via a uterine artery ipsilateral to the pregnant uterine horn at 2-hourly intervals. Utero-ovarian vein concentrations of 13,14-dihydro-15-keto PGF2 alpha (PGFM) and PGE2 were determined before and during oxytocin stimulation. Basal concentrations of both PGFM and PGE2 were significantly (P < 0.001) increased in ewes 15 h after RU486 administration compared with ewes receiving vehicle alone. Concentrations of PGFM, but not PGE2, increased significantly (P < 0.001) following injection of each dose of oxytocin in both treated and untreated animals. The response to oxytocin, measured both as the area under the curve and as the peak height of PGFM release, was significantly (P < 0.05) greater in RU486-treated ewes. There was no significant effect of oxytocin on the area or peak height of PGE2 response in either RU486-treated or control animals. These results demonstrate that treatment of late-pregnant ewes with RU486 results in an increase in basal uterine PGFM and PGE2 as well as oxytocin-stimulated PGFM release.


Regulation of female brain aromatase activity during the reproductive cycle of the dove. Oestrogen is formed in the female dove brain. The aim of this study was to determine whether (a) the catalytic properties of the brain aromatase are similar to the ovarian enzyme and (b) aromatase activity in the female brain changes during the reproductive cycle and is influenced by steroids and environmental stimuli. The results show that female preoptic aromatase has a higher substrate affinity than the enzyme in ovarian follicles (apparent Km: preoptic area, 7 nmol/l; ovarian follicles, 29 nmol/l), but a lower activity in the preoptic area (Vmax: preoptic area, 290 fmol/mg tissue per h; ovarian follicles, 843 fmol/mg tissue per h). In intact females with developing follicles, oestradiol-17 beta formation was higher in the posterior hypothalamus than the preoptic area. Females in a later stage of reproductive development (yolked follicles) had a different distribution of oestrogen formation with increased aromatase activity in the preoptic area. Preoptic and posterior hypothalamic aromatase activity of females paired with males for 10 days was positively correlated (r = 0.84, P = 0.0001; r = 0.75, P = 0.001 respectively) with ovarian development. Females with undeveloped ovaries which interacted with males had higher preoptic aromatase activity than visually isolated females with similar ovarian development, suggesting that behavioural stimuli have direct effects on brain aromatase activity which are independent of the ovary. Oestradiol benzoate treatment increased preoptic and posterior hypothalamic aromatase activity in intact and ovariectomized females, and testosterone propionate treatment increased anterior hypothalamic aromatase activity, but did not affect other areas, indicating that the distribution of induced aromatase activity is steroid-specific. Oestrogen treatment in ovariectomized or intact females did not replicate the maximal hypothalamic aromatase activity seen when the ovary contained yolked follicles. We conclude that brain aromatase activity is related directly to ovarian condition during the reproductive cycle of the female dove. As in the male, steroids have a role in the regulation of oestrogen formation in the female hypothalamus; behavioural stimuli are also likely to be involved in the control of the brain enzyme.


Immunological detection of the oestradiol receptor protein in cell lines derived from the lymphatic system and the haematopoietic system: variability of specific hormone binding in vitro. Extracts of human MCF 7 mammary carcinoma cells, the human lymphoblastoid cell lines AEH 1 and IM 9, T-cell derived CCRF cells, HL 60 myeloic leukaemia cells and murine myeloma cells SP 0 and NS I were analysed for immunoreactivity with polyclonal goat antibodies raised against homogeneous preparations of C-terminal fragments (32 kDa) of porcine uterine oestradiol receptor (ER). Whole cells and low speed cytosols were analysed for specific oestradiol-binding activity. ERs were enriched from cell extracts by either fractionated ethanol precipitation (0-25% (v/v) ethanol) and/or microscale-immunoaffinity chromatography. Immunoreactive proteins of identical molecular weight (approximately 65 kDa) were detected in all cell lines examined. Whole cell binding assays showed specific oestradiol-binding activity in MCF 7, IM 9 and CCRF cells. Borderline binding was found in HL 60 myeloid cells. No specific binding could be detected in AEH 1, NS I and SP 0 cells. Identical results were obtained using agar-electrophoresis after dextran-coated charcoal treatment. Immunoaffinity purified ERs from MCF 7, AEH 1 and HL 60 cells were subjected to limited proteolysis, where identical tryptic fragments were generated. In conclusion, we have confirmed by immunological methods that ERs are expressed in a variety of cell lines derived from the immune system and the haematopoietic system. The lack of specific hormone binding or very low-affinity hormone binding in some of the cells examined may be due to post-translational events or point mutations.


Sex hormone concentrations in blood serum from the north Atlantic fin whale (Balaenoptera physalus). Blood serum concentrations of testosterone and progesterone were measured in postmortem samples taken at sea from 814 fin whales (Balaenoptera physalus) caught during the summers (June-September) of 1981-1989. The ages of 781 of these animals were also assessed. The testosterone concentrations in samples from 352 males averaged 2 nmol/l; 41 samples had concentrations of 0.1 nmol/l or lower and 34 of these came from whales aged between 2 and 14 years and showed a Gaussian type of age distribution with a peak number at 7 to 8 years. The mean testosterone concentrations in the males increased by more than fourfold between June and August. Serum progesterone concentrations of the 462 females fell into three separate groups: (1) group I with values < or = 0.1 nmol/l; (2) group II with intermediate values of > 0.1 nmol/l but < 10 nmol/l; (3) group III with values of > or = 10 nmol/l. These three groups of females seemed to consist respectively of young sexually immature females, mature non-pregnant females and pregnant females. The age distribution in the groups indicated that puberty in females is attained chiefly between the ages of 7 and 10. The yearly pregnancy rate (that percentage of all females caught and studied in a year which had progesterone values > or = 10 nmol/l) was between 35% and 55%, except in 1987 when it was 67%. The yearly pregnancy rate would range from 56% to 93% if only mature females (i.e. those with serum progesterone > 0.1 nmol/l) were considered.(ABSTRACT TRUNCATED AT 250 WORDS)


Oestrogen enhancement of the myometrial response to exogenous parathyroid hormone-related protein (PTHrP), and tissue localization of endogenous PTHrP and its mRNA in the virgin rat uterus. Classical pharmacological studies have shown that oestrogen dominance in humans and other animals can increase the responsiveness of the uterus to many locally acting peptides. Parathyroid hormone-related protein (PTHrP) has been shown to be expressed in the pregnant and non-pregnant rat uterus and exogenous PTHrP is known to relax uterine contraction in vitro. We investigated whether oestrogen dominance can influence the responsiveness of the uterine horn to PTHrP, and further studied the localization of PTHrP mRNA and protein in the rat uterine horn using in-situ hybridization and immunohistochemistry. Exogenous PTHrP(1-34) inhibited spontaneous and electrically induced contractions in uteri isolated from non-cycling rats. Pretreatment of non-cycling rats with oestradiol-17 beta increased uterine sensitivity to PTHrP: EC50 values for inhibition of spontaneous contractions by PTHrP were 0.33 nmol/l, 1.1 nmol/l, 2.6 nmol/l and 7800 nmol/l in uteri from animals treated for 2 days with oestradiol-17 beta alone, 2 days with oestradiol-17 beta + 1 day progesterone, 1 day with oestradiol-17 beta alone and in untreated rats respectively. Similar EC50 values were obtained for electrically stimulated uteri. In agreement with these findings, uterine horns from cycling rats in pro-oestrous and oestrous phases of the cycle showed a higher responsiveness to PTHrP(1-34) when compared with uterine horns taken from rats in metoestrus and dioestrus. PTHrP mRNA and protein were detected in the endometrial epithelium lining of the lumen and the endometrial glands, as well as in the myometrium of rats which were either pretreated for 2 days with oestradiol-17 beta or untreated. This study suggests that PTHrP may act in an autocrine and/or paracrine manner to modulate uterine motility and function.


The effect of gonadotrophin surge-inhibiting factor on the self-priming action of gonadotrophin-releasing hormone in female rats in vitro. The present study was designed to explore further the functional antagonism between gonadotrophin-releasing hormone (GnRH) and the ovarian factor, gonadotrophin surge-inhibiting factor (GnSIF). In all experiments, pituitary tissue was exposed to various amounts of GnSIF, after which the self-priming action of GnRH was studied. GnSIF was increased in vivo by FSH treatment and increased in vitro by adding various amounts of follicular fluid (FF) to cultured pituitary cells. Treatment with 3 or 10 IU FSH suppressed the initial LH response and delayed the maximally primed LH response to GnRH. Treatment with FSH was only effective in intact rats on days 1 and 2 of dioestrus. There was no difference in the rate of maximal LH release irrespective of treatment with either FSH or saline. Since FSH treatment was ineffective in long-term ovariectomized rats, it was concluded that the initial suppressive effect of FSH on LH release was mediated by GnSIF. Cycloheximide prevented the self-priming action of GnRH by inhibiting GnRH-induced protein synthesis. The initial protein synthesis-independent GnRH-stimulated LH release, which was already suppressed by FSH treatment, remained suppressed in the presence of cycloheximide. Pretreatment with GnRH in vivo increased the protein synthesis-independent GnRH-induced LH release during subsequent incubation of the glands. This increase did not occur after FSH treatment. Pituitary cells, cultured for 20 h in medium only, failed to elicit the self-priming effect of GnRH. Preincubation with FF maintained the self-priming effect. This was independent of the concomitant presence of various amounts of oestradiol. Preincubation with bovine FF suppressed the initial GnRH-stimulated LH release dose-dependently. Porcine FF, human FF and testicular extract suppressed the release of LH in a similar way. It was concluded that GnSIF suppresses the initial LH response to continuous GnRH stimulation. Increased levels of GnSIF caused by FSH treatment also delayed the primed LH release. The mechanism of functional antagonism between GnSIF and GnRH could give rise to the occurrence of the phenomenon of GnRH self-priming.


Effects of placing micro-implants of melatonin in the mediobasal hypothalamus and preoptic area on the secretion of prolactin and beta-endorphin in rams. In a previous study, we showed that the local administration of melatonin in the mediobasal hypothalamus (MBH), but not the preoptic area (POA), caused a premature increase in the secretion of FSH and growth of the testes in sexually inactive Soay rams exposed to long days. To extend these observations, we have now measured blood concentrations of prolactin and beta-endorphin and the associated peripheral responses in the same animals, to establish whether the treatments produced multiple endocrine changes such as those which occur following exposure to short days. Groups of rams were initially exposed to alternating 16 weekly periods of long days (16 h light: 8 h darkness; 16L:8D) and short days (8L:16D) for at least 9 months to entrain the seasonal cycles in the secretion of the pituitary hormones. The treatments were started at 10 weeks under long days, when the animals had a physiology characteristic of the early summer with high blood plasma concentrations of prolactin (associated with growth of the summer pelage), and low concentrations of beta-endorphin (associated with low body weight). The animals were assigned at random to the following treatments: (i) micro-implants of melatonin in the MBH, (ii) microimplants of melatonin in the POA, (iii) empty implants in the MBH or POA to act as operated controls, and (iv) no surgery to act as unoperated controls (n = 12 rams/treatment). The micro-implants consisted of 22-gauge stainless-steel needles with melatonin fused inside the tip. The implants were inserted bilaterally in the brain, and left in place for 12-14 weeks. The observations continued for a total of 28 weeks while the animals remained under long days. The administration of melatonin in the MBH induced a rapid decreased in plasma concentrations of prolactin while in the POA it induced a less marked but significant effect. The mean times to minimum concentrations of prolactin were 7.4 +/- 0.4, 17.3 +/- 2.8 and 26.0 +/- 0.3 weeks for the MBH, POA and combined control groups respectively (MBH vs control, P < 0.001, POA vs control P < 0.01). In the MBH group, the concentrations of prolactin subsequently increased to a maximum 6 weeks after the end of melatonin treatment. The changes in prolactin were accompanied by changes in growth and moulting of the pelage; only animals in the MBH group showed a conspicuous moult associated with the change from low to high prolactin secretion.(ABSTRACT TRUNCATED AT 400 WORDS)


A simple bioassay for epidermal growth factor using pig thyrocytes. A bioassay for epidermal growth factor (EGF) is described using an eluted stain assay (ESTA) of dehydrogenase activity in pig thyrocytes. Optimal responsiveness to EGF was obtained in confluent cultures of primary pig thyrocytes cultured with EGF or biological samples containing EGF for either 24 or 48 h. Dehydrogenase activity was determined by measuring the production and then the release of a coloured formazan product produced by reduction of a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide substrate added to the cells. The assay responded equally to mouse, human and recombinant EGF and was suitable for measuring EGF activity in some but not all biological fluids. Specificity of detection of EGF activity was confirmed using antibody to EGF. The ESTA assay compared favourably with the radioreceptor assay for EGF in terms of sensitivity to EGF with half-maximal activation at 0.24 +/- 0.06 nmol/l (mean +/- S.E.M., n = 22 experiments) for the ESTA assay and 0.60 +/- 0.13 nmol/l (n = 7 experiments) for the radioreceptor assay.


The International Standard for Recombinant DNA-derived Erythropoietin: collaborative study of four recombinant DNA-derived erythropoietins and two highly purified human urinary erythropoietins. The International Standard (IS) for Recombinant DNA-Derived (rDNA) Erythropoietin (EPO) (in ampoules coded 87/684) and three other rDNA EPO preparations in ampoules coded 87/690, 87/696 and 88/574 respectively, were compared with two preparations of highly purified human urinary (HU) EPO and the 2nd International Reference Preparation of Human Urinary Erythropoietin for Bioassay (2nd IRP) by 26 laboratories in 11 countries using a wide range of in-vivo and in-vitro bioassays and immunoassays. These EPO preparations were also compared by electrophoresis and isoelectric focusing. Estimates of EPO content in terms of the 2nd IRP by all in-vivo bioassay methods gave combined unweighted geometric means (with 95% fiducial limits) of: 86 (75-99) IU/ampoule for the IS, 81 (70-94) IU/ampoule for 87/690, 58 (48-71) IU/ampoule for 87/696 and 120 (100-143) IU/ampoule for 88/574. Mean estimates of EPO content in terms of the 2nd IRP by in-vitro bioassays (except receptor assays) were larger than, and those by immunoassays were similar to, the mean estimates by in-vivo bioassays. The use of purified rDNA or HU EPO as standards in place of the 2nd IRP reduced the inter-laboratory variability of estimates of purified EPO preparations by in-vivo and in-vitro bioassays and by immunoassays, and reduced the variability of overall mean estimates for each of these preparations between the three types of method. The inter-laboratory variability of immunoassay estimates of human serum EPO was similar whether the 2nd IRP or one of the purified EPOs was used as standard. Significant differences in in-vivo and in-vitro biological, immunological and physicochemical properties were found between these four rDNA EPO preparations and between them and the HU EPO in the two purified preparations and in the 2nd IRP. There were also differences between the immunoreactivities of the two serum EPO samples included in the study, and between them and the immunoreactivities of the purified EPOs. The differences between rDNA EPOs appeared to be related to differences between the cells used for their biosynthesis, but may also be the result of differences in purification methods and of inter-batch variations. Significant differences in assay specificity were observed within each of the three general types of method. The specificity of the in-vivo bioassays was influenced by the route of hormone administration. The specificities of the mouse spleen cell in-vitro bioassays differed from that of the mouse spleen receptor-binding assay.(ABSTRACT TRUNCATED AT 400 WORDS)


The effects of systemic insulin, insulin-like growth factor-I and growth hormone on bone growth and turnover in spontaneously diabetic BB rats. Spontaneously diabetic BB rats have a markedly depressed longitudinal bone growth and bone formation/turnover. In this study, male diabetic BB rats were infused intraperitoneally or subcutaneously for 2 weeks with hormones that are believed to stimulate skeletal growth and/or trabecular bone formation: insulin (3 or 4 U/day), human GH (hGH; 400 mU/day), recombinant human insulin-like growth factor-I (rhIGF-I; 300 or 600 micrograms/day) and testosterone (80 micrograms/100 g body weight per day). Saline-treated diabetic BB rats had decreased plasma concentrations of IGF-I and osteocalcin (OC) (OC, 3.7 +/- 0.3 vs 13.1 +/- 0.8 (S.E.M.) nmol/l in controls); bone histomorphometry showed decreased epiphyseal width, osteoblast surface (0.04 +/- 0.04 vs 1.5 +/- 0.3%) and osteoid surface, and mineral apposition rate (MAR) (1.8 +/- 0.5 vs 7.9 +/- 0.6 microns/day). Testosterone and hGH infusions had no effect on weight loss or on decreased skeletal growth and bone formation of diabetic rats, nor did they increase plasma IGF-I concentrations. Insulin infusions into diabetic rats resulted in hyperinsulinaemia and accelerated weight gain. The epiphyseal width, osteoblast/osteoid surfaces and OC levels of insulin-treated rats were normalized or stimulated well above control values (osteoblast surface, 4.3 +/- 0.8%; plasma OC, 16.1 +/- 1.4 nmol/l); the MAR (4.0 +/- 0.9 microns/day) was only partly corrected after the 2-week infusion. Infusions of rhIGF-I into diabetic rats doubled but did not restore plasma IGF-I levels to normal; weight gain, however, was similar to that in control rats. IGF-I treatment had no effect on epiphyseal width, osteoblast/osteoid surfaces and OC concentrations, but improved the decreased MAR (4.6 +/- 1.2 microns/day).(ABSTRACT TRUNCATED AT 250 WORDS)


Primiparous rhesus monkey mothers are more sensitive to the nursing-induced inhibition of LH and ovarian steroid secretion. The duration of lactational infertility is prolonged significantly in adolescent, primiparous rhesus monkey (Macaca mulatta) mothers compared with adult, multiparous mothers. The present study examined the hypothesis that this parity/age difference in lactational infertility is due to a difference in the physiological responsiveness to nursing behaviour between adolescent and fully adult mothers and is not a consequence of differences in nursing behaviour, per se. At 22 weeks postpartum, mother-infant pairs were randomly assigned to one of four conditions: primiparous, nursing restricted (PR; n = 9); primiparous, nursing unrestricted (PU; n = 11); multiparous, nursing restricted (MR; n = 12); and multiparous, nursing unrestricted (MU; n = 8). Nursing was restricted for a 2-week period by mothers wearing a primate vest which prevented suckling behaviour but allowed infants to interact with their mothers. Nursing restriction resulted in a significant increase in serum oestradiol concentrations in both PR and MR mothers. Although nursing bout frequencies and durations were similar between PU and MU mothers, serum oestradiol also rose in MU mothers but remained suppressed in PU mothers. Once the nursing manipulation period ended and all mothers were allowed to nurse ad libitum, serum oestradiol concentrations continued to rise in all but the PU mothers. This brief interruption of nursing at 22 weeks postpartum advanced the subsequent timing of the first postpartum ovulation in MR and PR mothers relative to that of PU mothers. Again, despite similarities in nursing behaviour, the occurrence of first ovulation was also advanced in MU mothers compared with PU mothers. Just prior to the first postpartum ovulation, females were randomly assigned to one of four treatment groups to determine the effects of nursing behaviour on the hormonal parameters of the luteal phase: primiparous, nursing restricted (PRL; n = 9); primiparous, nursing unrestricted (PUL; n = 11); multiparous, nursing restricted (MRL; n = 10); and multiparous, nursing unrestricted (MUL; n = 10). Nursing restriction significantly elevated serum progesterone concentrations in PRL females compared with other mothers. Serum concentrations of oestradiol were higher in PRL, MRL and MUL mothers relative to PUL females. Again, this difference in oestradiol between PUL and MUL mothers occurred despite similarities in nursing behaviour. These data suggest that parity/age differences in the period of lactational infertility are not due to differences in nursing behaviour but rather to an increased sensitivity to the inhibitory aspects of the suckling stimulus in adolescent primiparous mothers.(ABSTRACT TRUNCATED AT 400 WORDS)


In vitro intracanal temperatures produced during warm lateral condensation of Gutta-percha. In vitro intracanal temperatures produced during the war lateral condensation of gutta-percha were measured using a computerized recording system that allowed repeated obturations of a root canal model. The obturations were performed using a Touch 'n Heat unit. Temperatures were recorded to an accuracy of a hundredth degree centigrade by 16 intracanal themocouples connected to the computerized measurement system. The highest intracanal temperature recorded was 114.51 degrees C at a power setting of 6, while the mean intracanal temperature increase above the average room temperature ranged from 8.18 to 65.05 degrees C. In addition, the spreader was not uniformly heated to the same temperature throughout its entire length. The hottest point on the spreader was located 5 mm from the tip.


Comparison of conventional root canal obturation techniques with Thermafil obturators. This study compared microleakage of Thermafil Obturators to the single cone technique in Lexan blocks simulating root canals (n = 20) stained with Prussian blue dye. The microleakage of Thermafil and laterally condensed extracted human teeth was also investigated after the teeth were stained with India ink or Prussian blue dye over 1- or 2-wk dye immersion periods. In Lexan blocks there was no statistical difference in mean leakage between the single cone technique and Thermafil. Teeth with Thermafil Obturators exhibited greater leakage than those with the lateral condensation. Greater leakage was also seen where India ink was used. Dye immersion time did not have a significant influence on leakage scores. The combination of India ink with Thermafil demonstrated the greatest average leakage. This study demonstrated that Lexan canal simulating blocks do not mimic extracted human teeth for evaluating microleakage.


Physical dimensions, torsional performance, and metallurgical properties of rotary endodontic instruments. 3. Peeso drills. A laboratory study was performed on Peeso drills to determine their physical dimensions, torsional performance, and metallurgical properties. Samples were measured from each of sizes #1 to #6 of Peeso drills (type P) and sizes #009 to #023 of Peeso drills (type B-1) from the two manufacturers that distribute these instruments in the United States. They were also tested in clockwise and counterclockwise torsion using a digital torque meter instrument. It was not possible to evaluate completely some type P drills of size #4 and larger or type B-1 drills of size #016 and larger because the torsional moments exceeded the capacity of the test instrument. Scanning electron microscopic examination confirmed visual observations that the stainless steel Peeso drills exhibited ductile torsional fracture, in contrast to the carbon steel Peeso drills which fractured in a relatively brittle manner. The carbon steel Peeso drills exhibited a much greater tendency for torsional fractures in the bur head, along with considerably smaller values of angular deflection at separation compared to the stainless steel Peeso drills. This study is part of a continuing investigation to establish standards for all rotary endodontic instruments.


A radiographic comparison of two root canal instrumentation techniques. The ability of two instrumentation techniques to negotiate and enlarge small curved canals was com-compared radiographically. Fifty canals in extracted human molar teeth were instrumented by the progressive enlargement (PE) technique, a form of step-back preparation using standard K files, or by the balanced force (BF) technique using K files whose tips had been specially modified. By using drawings and projected radiographic images of the files, the position of the largest file used in the apical preparation, #30 or #35 for the PE technique and #45 for the BF technique, was compared with the position of a small file placed in the canal before instrumentation. The PE and BF techniques were equally capable of instrumenting small curved canals to their respective largest apical preparation sizes. However, at sizes equivalent to the largest apical preparation sizes used in the PE technique, the BF technique produced significantly less deviation from the center of the original canal.


Comparison of apical leakage in teeth obturated with a polyamide varnish or zinc oxide and eugenol cement using lateral condensation. The sealing ability of copal varnish has been investigated when used as an adjunct in restorative procedures, retrofilling procedures, and dontic obturation procedures. Barrier, a polyamide-type polymer cavity varnish, has been compared with copal varnishes in restorative dentistry. This investigation compared the apical sealing properties of a zinc oxide and eugenol sealer (Roth) with a polyamide varnish (Barrier). Twenty-four teeth were instrumented and divided into two groups, one obturated with gutta-percha and Roth sealer and one with gutta-percha and Barrier, using submersion in methylene blue dye to demonstrate apical leakage. Roth sealer exhibited less dye penetration than Barrier when used as an endodontic sealant (p < 0.02).


Antibodies in normal and diseased pulps reactive with microorganisms isolated from deep caries. Immunoglobulin molecules in the supernatant fluids (SF) from pulpal explant cultures have been observed to react with microorganisms implicated in infections of root canals. In this study, the reactivity of immunoglobulin molecules in the SF from normal and irreversible pulpitis pulps to six strains of predominant microorganisms isolated from the immediate layer of carious lesions above the pulps used for explant cultures was investigated using an enzyme-linked immunosorbent assay. Two ATCC strains of Eubacterium were also included in this assay. Specific antibodies to Lactobacillus casei subsp. casei, Lactobacillus casei subsp. rhamnosus, Lactobacillus acidophilus (I), (II), Streptococcus mutans, Bacteroides intermedius, Eubacterium brachy, and Eubacterium alactolyticum in the SF from the normal and irreversible pulpitis tissues were observed with a large variation of antibody levels in both groups. Immunodiffusion assays of the SF revealed that IgG was the major class of immunoglobulin in the normal as well as the irreversible groups. The presence of natural antibodies in the normal pulps suggested a possible protective role of antibodies during the invasive process of caries.


Effect of noncutting tipped instruments on the quality of root canal preparation using a modified double-flared technique. Fifty-one extracted human first molar teeth with intact crowns and mature root apices were divided into three groups. Root curvature was determined. One of the root canals in the mesial root of lower molars, or the mesiobuccal root in maxillary molars, was prepared in one of three ways. In group 1 the root canals were instrumented using a modified double-flared technique with noncutting tipped files (Flex R); in group 2 the same files were used with a step-back technique. Group 3 was prepared with conventionally tipped files (K-Flex) using the step-back technique. A low viscosity polyvinyl siloxane impression material was injected into the prepared root canals and the specimens were decalcified, dehydrated, and cleared. The preparation was evaluated subjectively according to various desirable or undesirable criteria. A rating for overall quality of preparation was given. Statistical analysis showed that the teeth in group 1 had better overall preparation than those in group 3 (p < 0.05). There were no other statistically significant differences between the groups. The mean time required for each preparation technique was not statistically significantly different (p > 0.1). The use of a modified double-flared technique with non-cutting tipped files was shown to be an effective method for the preparation of curved root canals.


Use of dental instruments for bridging during electric pulp testing. It has been suggested that dental instruments can be used as bridging instruments to facilitate electric pulp testing of teeth with extensive restorations. This study reports a clinical investigation to evaluate the effectiveness of this procedure. One hundred seventeen vital teeth in 20 volunteers were tested. Ten endodontically treated teeth functioned as controls. Following appropriate isolation and asepsis technique, baseline recordings of the threshold response with the electric pulp tester were taken. With the use of dental explorers and endodontic files to bridge between the probe tip and the tooth surface, recordings were made of the threshold responses. Findings indicate that electrically conductive dental instruments can be reliably used as bridging instruments with the electric pulp tester.


Root end isolation for retrograde fillings. Isolation and moisture control of the root end of a tooth with a large periapical lesion is normally accomplished with the aid of bone wax, cotton pellets, or sterile gauze saturated with a suitable hemostatic agent. An alternative procedure, a simple and effective rubber dam technique, is presented demonstrating isolation of an apex before reverse filling.


Thermal analysis of commercial Gutta-percha using differential scanning calorimeter and dynamic mechanical thermal analysis. Gutta-percha is the most widespread root canal filling material and knowledge of its chemical and thermomechanical properties is of a great interest to its use in clinical dentistry. In the present investigation, the results from differential scanning calorimeter were compared with those obtained by dynamic mechanical thermal analysis. A significant correlation was established between these results and the chemical composition of six different commercial samples of gutta-percha.


Effect of intracoronal bleaching on external cervical root resorption. The purpose of this study was to examine histologically and radiographically the effect of a bleaching agent, a mixture of sodium perborate and superoxol (30% hydrogen peroxide), without a heat source on the cervical root surface and periodontal tissue of endodontically treated teeth of dogs. Endodontic treatment was performed on anterior teeth of dogs. The endodontically treated teeth were then divided into two groups. One group received walking bleach procedures, while the other group did not. The animals were killed at intervals of 1 and 3 months after bleaching for histological and radiographic examination. External cervical root resorption was only observed in bleached teeth at 3-month but not at 1-month observation.


Autonomic nerve endings in the odontoblast/predentin border and predentin of the canine teeth of dogs. Autonomic nerve endings in the odontoblast/predentin border and the predentin were observed by electron microscopy in the canine teeth of dogs. Adrenergic nerve endings with a number of very electron-dense granular vesicles and a minority of agranular vesicles of different sizes were clearly observed, but seen rarely, at the odontoblast/predentin border, in the predentin adjacent to the odontoblast processes, and as free endings in the middle part of the predentin. Cholinergic nerve endings were also observed with a number of agranular vesicles of different sizes without any visible exocytosis, in the odontoblast/predentin border, and in the predentin. Near the areas of collagen formation, extensive exocytosis of granular vesicles of different sizes was observed in adrenergic nerve endings located as free nerve endings in the predentin.


A bacteriological and histological evaluation of 58 periapical lesions. Periapical tissue from 58 cases requiring periapical surgery was examined histologically and cultured for the presence of microbes. Twenty-nine had a possible oral cavity communication and 29 did not. Approximately one-half of each biopsy was submitted for culture while the other portion was examined histologically. Cultures were positive for the presence of bacteria in 51 of 58 cases while bacteria were seen histologically in only 8 of 58 cases. A total of 50 different species of bacteria were isolated from the 58 cultures of periapical tissue. Of 133 isolates, 87 were strict anaerobes, 37 were facultative anaerobes, and 9 were aerobes. Bacteroides species were found in 17 cultures, always with additional bacteria. Seventeen of 58 biopsies contained foreign particulate matter thought to be root canal sealer. Bacteria were found in periapical granulomas, radicular cysts, and a periapical abscess. According to our data, bacteria, foreign material, missed canals, vertical root fractures, and periodontal disease may all contribute to the chronic, non-healing periradicular lesion.


A comparison of apical seal: chloroform versus halothane-dipped gutta-percha cones. A dye penetration study was done to compare apical leakage among three groups of extracted teeth obturated with a lateral condensation technique. In one group the master gutta-percha cones were customized in the apical portion of the canals after being dipped in chloroform. A second group used halothane as the customizing agent. No dip was used in the third group, and all three were then laterally condensed. The teeth were cleared and dye penetration was measured. Statistical analysis using a Kruskal-Wallis one-way analysis of variance of the data showed no significant difference among the groups at the p < 0.05 level.


Anatomical relationship of the mandibular canal to its surrounding structures in mature mandibles. A lack of agreement exists in the literature regarding the anatomical relationship of the mandibular canal to its surrounding structures such as the root apices. The purpose of this investigation was to study the spatial relationship of the mandibular canal to the posterior teeth in dried human mandibles. Twenty-two mature dried mandibles were sectioned through the root apices of the first and second premolars and molars. Second premolars and second molars had the closest distances to the canal with a mean of 4.7 mm and 3.7 mm, respectively. With a mean of 6.9 mm, the apices of the mesial roots of the first molars were farthest from the canal. The canal pathway in mature mandibles followed in S-shaped curve in 31% of the cases. In 41% of the cases it was located lingual (19%), buccal (17%), or directly inferior (5%) to the apices of the posterior teeth. In 28% of the cases the canal could not be identified clearly in the second premolar and first molar regions. In a typical S-shaped configuration the canal was located buccal to the distal root of the second molar, crossed to the lingual below the second molar mesial root, ran lingual to the first molar, and crossed back to the buccal apical to the apex of the second premolar. Based on our results it appears that the mandibular second premolar and second molar are the most likely teeth to be involved in accidental damage to the mandibular canal during root canal therapy.


Endodontic interappointment flare-ups: a prospective study of incidence and related factors. Severe pain and/or swelling following a root canal treatment appointment are serious sequelae. Information varies or is incomplete as to the incidence of these conditions and related factors. In this study, data were collected at root canal treatment appointments on demographics, pulp/periapical diagnoses, presenting symptoms, treatment procedures, and number of appointments. Patients that then experienced a flare-up (a severe problem requiring an unscheduled visit and treatment) had the correlating factors examined. Statistical determinations were by chi-square analysis with significance at 0.05 or less. Nine hundred forty-six visits resulted in an incidence of 3.17% flare-ups. Flare-ups were positively correlated with more severe presenting symptoms, pulp necrosis with painful apical pathosis, and patients on analgesics. Fewer flare-ups occurred in undergraduate patients and following obturation procedures. There was no correlation between patient demographics or systemic conditions, number of appointments, treatment procedures, or taking antibiotics.


Healing of a sinus tract of periodontal origin. A sinus tract adjacent to teeth is usually considered to be of endodontic origin and root canal therapy is the primary treatment to achieve healing. A case of a sinus tract is presented in which a suspected endodontic-periodontic lesion was successfully treated by periodontal therapy alone. The sinus tract healed and the radiograph showed bony repair.


Effects of moisture content and endodontic treatment on some mechanical properties of human dentin. The objective of this study was to determine whether significant differences exist between the mechanical properties of human dentin from treated pulpless teeth and dentin from normal vital teeth. Dentin specimens (n = 262) were obtained from 54 freshly extracted normal vital human teeth and 24 treated human pulpless teeth. These specimens were subjected to different experimental conditions (wet, air dried, desiccated, and rehydrated). Compression, indirect tensile, and impact tests were conducted to measure the mechanical properties of those specimens. All data obtained were analyzed with t tests. The results showed that the dehydration of dentin increases the Young's modulus, proportional limit (in compression), and especially the ultimate strength (in both compression and tension). Substantial dehydration changes the fracture characteristics of dentin specimens under static compressive and indirect tensile loadings. The measurements of impact-breaking energies of desiccated dentin were not found to be significantly decreased. The compressive and tensile strengths of dentin from treated pulpless teeth obtained in this study do not appear to be significantly different from those of normal dentin (p > 0.05), while the mean values of Young's modulus and proportional limit in compression tests appear to be lower. Fifty percent of the dentin specimens from treated pulpless teeth exhibit greater plastic deformation than normal dentin in compression. The results of this study do not support the theory that dehydration after endodontic treatment per se weakens dentin structure in terms of compressive and tensile strengths. Other mechanical properties of treated pulpless teeth, however, may not be the same as those of normal vital teeth.


Histological evaluation of the presence of bacteria in induced periapical lesions in monkeys. In endodontic periapical lesions, both presence and location of bacteria are controversial. Various experimental techniques have produced differing results perhaps related to potential artifacts such as contamination during specimen recovery. Our objective was to examine for bacteria in uncontaminated, undisturbed periapical lesions in an animal model. Pulp necrosis was induced by exposing molars in nonhuman primates and closing the exposure after 1 week with amalgam. Lesions developed at 18 apices. After 7 months, block sections including tooth and surrounding tissues were removed, processed histologically, and Gram stained. Bacteria, primarily Gram positive, were consistently identified in necrotic tissue in canals. Two canals demonstrated bacterial masses to the apical foramen. No bacterial colonies, only intracellular microorganisms, were seen periapically. Inflammatory lesions seemed to resist the spread of bacteria, confining them to the canal space. Bacterial masses at the apical foramen could contaminate periapical tissues during surgery or extraction and give a false positive upon microbiological sampling.


Torsional properties of the Canal Master instrument. The torsional properties of a new type of endodontic hand instrument, the Canal Master, were compared with conventional machined K-type endodontic files. Sizes 20 through 50 were tested in clockwise (CW) and counterclockwise (CCW) directions. Canal Master instruments exhibited significantly less torque at yield and at failure in both the CW and CCW directions. Rotation at failure was significantly greater for the Canal Master instruments in both CW and CCW directions. There was no significant difference in either torque or the amount of rotation for the Canal Master instruments when comparing CW versus CCW rotation. There was no clinically visible evidence of deformation of the Canal Master instruments prior to torsional failure.


Response of pulpal blood flow to intra-arterial infusion of endothelin. Laser Doppler flowmetry was used to determine the effect of the vasoconstrictor endothelin 1 (ET-1) on the pulpal blood flow of intact dogs' teeth during mandibular arterial infusion. ET-1 produced a profound decrease in pulpal blood flow of a relatively long duration at lower doses than similar infusions of norepinephrine. The decrease in pulpal blood flow in response to ET-1 was partially attenuated by the calcium channel blocker nifedipine. These findings demonstrate the presence of receptors for ET-1 in the microvasculature of the dental pulp and suggests that ET-1 may function in the local control of the pulpal microcirculation.


Effect of precurving on the performance of endosonic K files. This study compared the performance of precurved and straight endosonic files. Size 15, 20, and 25 endosonic K files were precurved to different degrees (20 to 90 degrees) and the resultant oscillatory pattern showed no significant difference compared with corresponding straight files. Endovue blocks were prepared with either a #15, 20, or 25 endosonic file which was either straight or precurved. Those blocks prepared with precurved files had a continuous taper while blocks prepared with straight files had constrictions along their lengths. Finally, a curved root of a human natural tooth had windows prepared along its length so that the oscillatory pattern of the file could be observed. The precurved file oscillated more freely than a straight file as observed by the presence of antinodes along the file and the accumulation of dentin chips within the canal. The results of this study suggest that it is advantageous to precurve endosonic files before using them in curved canals.


Magnetic resonance imaging of human teeth. There is a need in clinical dentistry for improved diagnostic methods for an accurate evaluation of pulp tissue pathosis. The purpose of this investigation was to attempt to image extracted human teeth with the aim of establishing the possible future application of magnetic resonance imaging in the diagnosis of pulpal pathosis. The results of this pilot study demonstrate that, in the in vitro setting, the outline of the tooth and pulp chamber can be imaged with a high degree of accuracy, and detailed images of the periodontal membrane and gross pulpal anatomy can be obtained. There were obvious differences in the signals given off by different locations within these teeth, and the images of the dental pulp tissue were detailed enough to suggest the possible correlation with histology. The high (9.4 T) field strengths used in this study demonstrate the in vitro application of this technology and the future possibilities for diagnosis of periodontal and odontogenic problems are significant.


Endodontic stabilizers.

Endodontic stabilizers. With the recent influx of osseointegrated endosseous implants in dentistry, attention has been drawn away from using endodontic stabilizers as a means of stabilizing and retaining teeth. It is our opinion that endodontic stabilizers should be considered in the treatment planning of seemingly nonretainable teeth. They are biocompatible and have the additional advantage of maintaining the periodontal membrane attachment of the remaining tooth. Extraction and subsequent replacement with osseointegrated implants should only be considered after all other means of retaining the natural tooth have been fully explored. Selective case studies from various areas of the dentition demonstrate the broad applicability of stabilizers. A technique is presented for the use of endodontic stabilizers.


Reference block for auxiliary cones.

Reference block for auxiliary cones. A technique is described for fabricating a reference block which will allow the practitioner to select the accessory cones most correctly corresponding to the spreader used. The technique utilizes materials common to the dental office and is simple enough to allow fabrication of a new block whenever the obturation instruments or materials are changed.


Effects of CO2 laser energy on dentin permeability. The effect of a CO2 laser on the structure and permeability of smear layer-covered human dentin was evaluated in vitro. Three different energy levels were used (11, 113, and 566 J/cm2). The lowest exposure to the laser energy increased dentin permeability, measured as a hydraulic conductance, due to partial measured as a hydraulic conductance, due to partial loss of the superficial smear layer and smear plugs. The intermediate energy level also increased dentin permeability by crater formation, making the dentin thinner. The lack of uniform glazing of the surface of the crater, leaving its surface porous and in communication with the underlying dentinal tubules also contributed to the increase in dentin permeability seen with the intermediate laser energy. The highest laser energy produced complete glazing of the crater surfaces and sealed the dentinal tubules beneath the crater. However, it also completely removed the smear layer in a halo zone about 100-microns wide around each crater which increased the permeability of the pericrater dentin at the same time it decreased the permeability of the dentin within the crater. The combined use of scanning electron microscopy and permeability measurements provides important complementary information that is essential in evaluating the effects of lasers on dentin.


Thermomechanical analysis of dental gutta-percha. Samples of three common commercial gutta-percha endodontic filling points (Hygienic, Mynol, and Maillefer Pink) and a sample of natural gutta-percha were submitted to thermomechanical analysis under pressures ranging from 0.01 to 0.2 N. Samples of Mynol and Maillefer filling points were thermally treated and then submitted to thermomechanical analysis in parallel with differential scanning calorimetric analysis. The results show that thermomechanical analysis gives results distinct from those obtained by classic dilatometry and that it is a technique well suited to the study of the thermoplastic properties of gutta-percha. An analysis of the results shows that the amount of the inorganic component used in a commercially available endodontic point has a strong influence on its thermomechanical properties. Thermodynamical properties and the "thermal history" of the gutta-percha are also important. Both temperature and force should be controlled in order to assess the thermomechanical properties of endodontic filling points, while the latter have not yet been codified in clinical procedures.


The mental foramen: 2. Radiographic position in relation to the mandibular second premolar. Seventy-five adult human mandibles were radiographed with a paralleling technique to determine the ability to visualize the mental foramen as well as its size and position. The foramen was seen on 75% of the horizontal periapical radiographs examined. When the foramen was not visualized, is was usually below the inferior edge of the film. The radiographic size of the foramen was smaller than the anatomical size previously reported. The position of the mental foramen was usually mesial and below the radiographic apex of the second premolar.


Assessment of external root resorption using digital subtraction radiography. Digital subtraction radiography was investigated for its capability to detect and quantify experimentally produced external root resorptive defects in teeth. Using a long source to object X-ray technique and E-speed film, serial radiographs of teeth with artificial lesions in a dry human skull (soft tissue simulated) were obtained. Receiver operating characteristic analysis was used to evaluate the diagnostic performance for each imaging system (conventional versus subtraction). To explore the quantitative assessment potential of digital subtraction radiography, images were produced after sequential demineralization by HCl. The acid solution was analyzed for calcium concentration by atomic absorption spectrophotometry. Three-dimensional histogram quantification for each subtracted image was performed. In overall performance for detecting experimentally produced external root resorption, digital subtraction radiography was found to be significantly superior to conventional radiography. In addition, digital subtraction radiography can provide quantification of experimentally produced external root resorptive defects.


Evaluation of Thermafil obturation of curved canals prepared by the Canal Master-U system. A dye penetration study was performed to compare the apical seals obtained with laterally condensed gutta-percha and Thermafil. The sample group contained 40 teeth with curved canals which had been prepared with the Canal Master-U System. Canal curvature was measured and the teeth were decalcified, cleared, and linear dye penetration was measured. Thermafil had statistically more linear leakage as verified by a Wilcoxon test (p < 0.001). Thermafil also had greater variability in the apical seal than did laterally condensed gutta-percha as verified by an F test (p < 0.001). A Spearman correlation analysis (p > 0.05) showed that there was no correlation between root curvature and linear dye penetration.


Effect of bleaching agents on inorganic components of human dentin and cementum. The effect of bleaching agents on the inorganic composition of human dentin and cementum was examined. Intact teeth were crushed, pulverized, and separated to dentin and cementum powders. The pulverized tissues were exposed to treatments with 30% H2O2, 3% H2O2, 2% sodium perborate in 30% H2O2, 2% sodium perborate in 3% H2O2, and 2% sodium perborate in bidistilled water for periods of 15 min and 1, 24, and 72 h. The degree of dissolution and the percentage of inorganic material for both dentin and cementum were measured. Thirty percent H2O2 and 2% sodium perborate in 30% H2O2 treatments significantly increased the solubility of dentin and cementum. The degree of dissolutions and the percentage of inorganic material remaining in the undissolved dentin and cementum increased with time progression. The greatest increase occurred with 30% H2O2 and 2% sodium perborate in 30% H2O2 after 24- and 72-h treatments. It is concluded that 30% hydrogen peroxide treatment may cause alteration in the chemical structure of the dentin and cementum making them more susceptible to degradation.


A three-dimensional study of canal curvatures in the mesial roots of mandibular molars. The degree and configuration of canal curvature was studied in the mesial roots of 100 randomly selected mandibular first and second molars. The teeth were radiographed in buccolingual (clinical) and mesiodistal (proximal) directions with #8 K files in place. One hundred percent of the specimens demonstrated curvature in both views. No correlation in degree of curvature was found to exist between the clinical and proximal views. Secondary curvature, in a direction opposite to that of the principle curve, was seen more frequently in the proximal view. In the proximal view, canals exhibited greater mean curvature than in the clinical view 38% of the time. Weine type II morphology (two canals, one foramen) demonstrated the greatest range in canal curvature when viewed from the proximal. Coronal flaring with Canal Master rotary instruments to a level just coronal to the curve significantly reduced the severity of curvatures in both views for most cases.


Chloroform in the endodontic operatory.

Chloroform in the endodontic operatory. This article reviews the role chloroform has played in dentistry and describes an occupational health clinical investigation into the possible hazards of chloroform use in the operatory. Due to a Food and Drug Administration ban on drugs and cosmetics containing chloroform, there has been some confusion as to whether the use of chloroform in the practice of dentistry is considered unsafe or has been prohibited. Utilizing common endodontic treatment methods employing chloroform, this study reports no negative health effects to the dentist or assistant and air vapor levels well below Occupational Health and Safety Administration mandated maximum levels. The report concludes that, with careful and controlled use, chloroform can be a useful adjunct in the practice of dentistry. The Food and Drug Administration has no jurisdiction over a dentist's use of chloroform in clinical practice and has not proven that chloroform is a human carcinogen.


Odontogenic cutaneous sinus tract.

Odontogenic cutaneous sinus tract. Patients with odontogenic cutaneous sinus tracts usually seek treatment from a physician instead of a dentist and the etiology is frequently misdiagnosed. This case report describes a patient who sought treatment from a dentist because of a history of a previous cutaneous swelling on the ipsilateral side of his cheek. The etiology of the present swelling was a carious tooth. The patient was treated concurrently with both nonsurgical endodontic therapy and transcutaneous drainage by aspiration. This method of drainage precluded possible spontaneous drainage and avoided damage to proximal anatomical structures which may have occurred if an incision had been used.


Penetration of the pulp chamber by carbamide peroxide bleaching agents. Hydrogen peroxide readily penetrates the pulp chamber of freshly extracted teeth. This study was undertaken to determine whether carbamide peroxide also penetrates the pulp chamber. Freshly extracted teeth were sectioned 2 to 3 mm apical to the cementoenamel junction and the coronal pulpal tissue was removed. Acetate buffer was placed in the pulp chamber to absorb and stabilize any peroxide that might penetrate. The coronal portion of each tooth was immersed in either carbamide peroxide gel or gelled hydrogen peroxide at various concentrations for 15 min at 37 degrees C. The buffer was removed, leukocrystal violet was added, and the optical density of the resulting blue solution was determined spectrophotometrically. Amounts of peroxide found in the pulp chamber after 15 min ranged from 3.3 +/- 0.38 micrograms for the 10% sample to 40.4 +/- 3.51 micrograms for the 30% sample.


Master cone apical behavior under in vitro compaction. This study first evaluated the fitting of two brands of gutta-percha points 0.5 and 1.0 mm short of the apical end of an artificial canal. Photographs were taken before and after vertical or lateral compaction and measurements were made. Warm gutta-percha compaction was associated with greater apical cone movement than lateral condensation. Total apical cone movement was not related to the distance the master cone was initially adapted from the apical opening.


Sealing ability of Thermafil with and without sealer. The purpose of this study was to evaluate the apical sealing ability of root canals filled using three obturating techniques. Sixteen maxillary first molars were obturated with Thermafil, 16 with Thermafil and a ZOE sealer, and 16 with laterally condensed gutta-percha. Two canal instrumentation methods were used, conventional step-back preparation with K-Flex files and traditional instrumentation combining reaming with reamers and filing with K-Flex files. Following obturation, the teeth were prepared for evaluation of the seal using India ink and a Profile Projector. The depth of penetration of the dye was statistically evaluated for each of the three roots with an analysis of variance. For sealing ability, there was no significant difference at the p < 0.05 level between the conventional step-back preparation and the traditional instrumentation technique. However, a significant difference was present at the p < 0.05 level for the obturation techniques. The mean linear dye penetration for the Thermafil technique was greater than that for lateral condensation.


In vivo and in vitro glycosaminoglycans from human dental pulp. A qualitative assessment was made of the type of glycosaminoglycans (GAG) present in normal human dental pulp using electrophoresis on cellulose-acetate plates. A comparison was also made between the GAG derived directly from the dental pulp (in vivo) and those derived from cultured pulp fibroblasts from the same individual (in vitro). The results of this study showed four main types of GAG in normal human dental pulp tissue, which were dermatan sulfate, heparan sulfate, hyaluronic acid, and chondroitin sulfate. GAG synthesis from cultured pulp fibroblasts in vitro was different from the GAG present in the dental pulp (in vivo). Extracellular GAG, as well as pericellular GAG consisted of dermatan sulfate, hyaluronic acid, chondroitin sulfate, and heparin. Cellular GAG, however, contained only dermatan sulfate, hyaluronic acid, and chondroitin sulfate. There was no difference in type of GAG from the second and fourth passaged pulp fibroblasts.


Are endodontically treated teeth more brittle? This study compared biomechanical properties (punch shear strength, toughness, hardness, and load to fracture) of 23 endodontically treated teeth (mean time since endodontic treatment: 10.1 yr) and their contralateral vital pairs. Analyses using paired t tests revealed no significant differences in punch shear strength, toughness, and load to fracture between the two groups. Vital dentin was 3.5% harder than dentin from contralateral endodontically treated teeth (p = 0.002). The similarity between the biomechanical properties of endodontically treated teeth and their contralateral vital pairs indicates that teeth do not become more brittle following endodontic treatment. Other factors may be more critical to failure of endodontically treated teeth.


Canal Master files: scanning electron microscopic evaluation of new instruments and their wear with clinical usage. Canal Master instruments have a short, fluted cutting area and a small flexible shaft; they may be predisposed to rapid wear and breakage. This study examined new and used files as to tip, flute, and shaft design when new and with increasing time of canal preparation. One hundred files prepared 140 curved canals. Sizes #20, #40, and #60 files were examined unused and after 1, 3, 5, and 7 min of use. Evaluation was for uniformity when new and for deterioration and breakage with usage. The smaller sizes (#20 and #40) had some inconsistencies when new and were most predisposed to wear and breakage with time. Findings suggest that smaller files could have had improved quality control by the manufacturer. Also, they should be used with caution and discarded after short times of use in small, curved canals. This should minimize instrument separation and maximize cutting efficiency.


An evaluation of the Thermafil endodontic obturation technique. The purpose of this study was to evaluate the Thermafil endodontic obturation technique and to compare it with laterally condensed gutta-percha. Thirty-seven maxillary central incisors were similarly prepared and divided into groups. Seventeen were obturated with Thermafil and 17 with lateral condensation. Three served as controls. After vacuum staining, all teeth were cleared and apical dye penetration was evaluated by two independent observers. Average leakage values were 0.24 mm and 1.32 mm for Thermafil and 0.47 mm and 1.18 mm for lateral condensation. There was no significant difference between the techniques, although a difference between evaluators was noted. Final results point to the relative subjectivity of in vitro leakage studies. When comparing the obturation times of both techniques, the Thermafil technique averaged 2 min 56 s while lateral condensation took 3 min 26 s. Statistical analysis showed no significant difference in obturation times.


Anatomical study of the root apex in the maxillary anterior teeth. The purpose of this study was to investigate anatomically the apical portion of the root canal of human maxillary anterior teeth. Thirty maxillary central incisors, 30 maxillary lateral incisors, and 30 maxillary cuspids were used. These were teeth from patients ranging in age from 11 to 73 yr. The root apex and main apical foramen coincided in 16.7% of central incisors and cuspids and in 6.7% of the lateral incisors. The labiolingual diameter of the root canal at the apical constriction of the central incisors, lateral incisors, and cuspids averaged 0.425 mm, 0.369 mm, and 0.375 mm, respectively; and the respective vertical distances between apex and apical constriction were 0.863 mm, 0.825 mm, and 1.010 mm.


The standardized stop: a new concept in endodontics. This article presents a technique for making and using disposable standardized silicone stops. The stops have several diameters, colors, and thicknesses in order to make nonsurgical endodontics more efficient and improve the quality of therapy.


Adverse response to vital bleaching.

Adverse response to vital bleaching. A case study is presented that described an acute flare-up of a tooth following a vital bleaching procedure. The case illustrates (a) the importance of assessing the pulpal and periradicular status of a tooth prior to any vital bleaching procedure; and (b) the alteration of the local adaptation syndrome by the bleaching agent.


Behavioral variability and frequency-dependent selection. In Experiment 1, two conditions were compared: (a) a variability schedule in which food reinforcement was delivered for the fourth peck in a sequence that differed from the preceding N four-peck sequences, with the value of N continuously adjusted to maintain reinforcement probability approximately constant; and (b) a control condition in which the variability constraint was dropped but reinforcement probability remained constant. Pigeons responded approximately randomly under the variability schedule but showed strong stereotyped behavior under the control condition. Experiments 2 and 3 tested the idea that variability is the outcome of a type of frequency-dependent selection, namely differential reinforcement of infrequent behavior patterns. The results showed that pigeons alternate when frequency-dependent selection is applied to single pecks because alternation is an easy-to-learn stable pattern that satisfies the frequency-dependent condition. Nevertheless, 2 of 4 pigeons showed random behavior when frequency-dependent selection was applied to two pecks, even though double alternation is a permissible and stable stereotype under these conditions. It appears that random behavior results when pigeons are unable to acquire the stable stereotyped behavior under a given frequency-dependent schedule.


Key pecking of pigeons under variable-interval schedules of briefly signaled delayed reinforcement: effects of variable-interval value. Key pecking of 4 pigeons was maintained under a multiple variable-interval 20-s variable-interval 120-s schedule of food reinforcement. When rates of key pecking were stable, a 5-s unsignaled, nonresetting delay to reinforcement separated the first peck after an interval elapsed from reinforcement in both components. Rates of pecking decreased substantially in both components. When rates were stable, the situation was changed such that the peck that began the 5-s delay also changed the color of the keylight for 0.5 s (i.e., the delay was briefly signaled). Rates increased to near-immediate reinforcement levels. In subsequent conditions, delays of 10 and 20 s, still briefly signaled, were tested. Although rates of key pecking during the component with the variable-interval 120-s schedule did not change appreciably across conditions, rates during the variable-interval 20-s component decreased greatly in 1 pigeon at the 10-s delay and decreased in all pigeons at the 20-s delay. In a control condition, the variable-interval 20-s schedule with 20-s delays was changed to a variable-interval 35-s schedule with 5-s delays, thus equating nominal rates of reinforcement. Rates of pecking increased to baseline levels. Rates of pecking, then, depended on the value of the briefly signaled delay relative to the programmed interfood times, rather than on the absolute delay value. These results are discussed in terms of similar findings in the literature on conditioned reinforcement, delayed matching to sample, and classical conditioning.


Competition between stimulus-reinforcer contingencies and anticipatory contrast. Procedures used to study anticipatory contrast are conceptually similar to those used to study autoshaping, in that two target stimuli signal either higher or lower rates of reinforcement in the following components of the schedule. Despite this signal contingency, anticipatory contrast entails response rates that are higher to the target stimulus followed by the lower rate of reinforcement. To determine the relation between such effects and autoshaping, different variations of the procedure were used in which the signal contingency was presented in the absence of reinforcement in the target components themselves and in which the reinforcement schedules in the different following components were signaled by the same stimulus. Autoshaping effects of this signal contingency were demonstrated when no reinforcement was available during the target-component signals themselves. Intermediate patterns of behavior occurred when reinforcement was available during the target-component signals and when their different following schedules were correlated with the same stimulus. Attempts to isolate these signal and contrast effects functionally by using the signal-key procedure were unsuccessful. The results demonstrate that Pavlovian stimulus contingencies are in competition with the dynamics of anticipatory contrast, thus reducing its occurrence under some circumstances.


Successive independence and behavioral contrast in a closed economy. Two pigeons had access to multiple concurrent schedules of reinforcement for 24 hours per day in their home cages. The variable-interval schedules comprising the multiple concurrent schedules were varied across 16 conditions. In three sets of conditions, one schedule was varied while its concurrent alternative and the concurrent schedules in the other component were held constant. Behavioral contrast was observed; that is, as the rate of reinforcement arranged by the varied schedule decreased, response rates on the constant schedules typically increased. These conditions formed part of two larger sets of conditions in which the concurrent schedules in one multiple-schedule component remained constant while the concurrent schedules in the other component were varied. Successive independence was found, in that behavior allocation during the constant component did not vary as a function of the reinforcer ratios in the varied component. Successive independence between components in multiple concurrent schedules is a robust result that occurs in closed economies and under conditions that promote behavioral contrast.


Conjoint schedules of timeout deletion in pigeons. This experiment attempted to bring behavior under joint control of two distinct contingencies, one that provided food and a second that extended the periods during which that food was available. Pigeons' responses on each of two keys were reinforced according to a single random-interval schedule of food presentation except during signaled timeout periods during which the schedule was temporarily disabled. By means of a conjoint schedule, responses on the initially less preferred key not only produced food but also canceled impending timeouts. When behavior came to predominate on this conjoint alternative, the consequences of responding on the two keys were reversed. Responding in 3 of 4 pigeons proved sensitive to the conjoint scheduled consequences, as evidenced by systematic shifts in response rates favoring the conjoint key. In 2 of these 3 pigeons, sensitivity to the conjoint contingency was evident under time-in:timeout ratios of 2:1 (time-in = 120 s, timeout = 60 s) and 1:5 (time-in = 30 s, timeout = 150 s), whereas for the other pigeon preference for the conjoint key was observed only under the latter sequence of conditions. There was only weak evidence of control by the conjoint scheduled consequences in the 4th subject, despite extended training and forced exposure to the conjoint alternative. The overall pattern of results is consistent with studies of timeout avoidance but also shares features in common with positively reinforced behavior.