Control of shell settling in the swimming sea anemone Stomphia coccinea. 1. Electrical activity has been recorded from Stomphia coccinea during the behavioural sequence in which the detached anemone settles on to a Modiolus shell. 2. When a responsive tentacle contacts the shell, a short, complex burst of pulses is elicited. These remain confined to the region of contact. The endodermal slow-conduction system (SS2) then begins to fire repetitively (a typical example is 16 SS2 pulses at a mean interpulse interval of 5 s) until the pedal disc begins to inflate. Shell-tentacle contact is essential for stimulation of SS2 activity. 3. The complete response, apart from local bending of the column, may be reproduced by electrical stimulation of the SS2 alone. As few as 10 stimuli at frequencies between 1 shock/s and 1 shock/10 s are required to elicit the response.


Two slow conduction systems co-ordinate shell-climbing behaviour in the sea anemone Calliactis parasitica. 1. Pulses in two slow conducting systems, the ectodermal SS 1 and the endodermal SS 2, were recorded during shell-climbing behaviour. The mean pulse interval of SS 1 pulses was 7-4 s and that of SS 2 pulses was 6-4 s. Activity in both systems may arise as a sensory response of tentacles to shell contact, but the SS 1 and SS 2 may not share the same receptors. 2. Electrical stimulation of the SS 1 and SS 2 together, at a frequency of 1 shock every 5 s, elicits shell-climbing behaviour in the absence of a shell. 3. Low-frequency nerve-net activity (about 1 pulse every 15 s) accompanies column bending during both normal and electrically elicited responses. This activity probably arises as a result of column bending and is not due to a sensory response to the shell.


Responses of trout gill ion transport systems to acute acidosis. The response of rainbow trout Na+ and Cl- uptake systems to acute acidosis was tested by slow infusion of lactic acid into anaesthetized animals. Depression of blood pH by 0-4 pH unit had no effect on influx rates for either ion, and we conclude that gill ion uptake systems do not respond rapidly to blood pH changes.


On the interaction of NH+4 and Na+ fluxes in the isolated trout gill. 1. Sodium influx was measured in isolated, previously perfused gill arches of rainbow trout, Salmo gairdneri, by measuring incorporation of 22Na into gill tissue following timed exposure to a 1 mM 22NaCl medium. Transport rates approximated those estimated for intact fish and were linear for at least one min. 2. NH4Cl-containing perfusates at pH 7 and 8 stimulated Na+ influx equally, indicating that only ionized ammonia is important in the transport process. A Na+/NH4+ exchange at basal and/or lateral membranes of the transporting cells is suggested. 3. Low-sodium Ringer perfusate augmented Na+ influx; in one group of gills the transport rate was more than double that of NaCl Ringer controls. The increase in transport induced by internal NH4+ was not additive with the low sodium augmentation. A reduction in intracellular (Na+) is postulated as the mechanism operating in both cases. 4. Ouabain had no appreciable effect on Na+ influx, either with or without NH4+ in the perfusate. Diamox partially blocked the augmented Na+ influx induced by NH4+. Amiloride completely inhibited Na+ influx, both with and without NH4+ in the perfusate.


The mobilization of calcium and bicarbonate by raised concentrations of potassium in the haemolymph of the snail, Helix pomatia. 1. After the concentration of potassium in the haemolymph of Helix pomatia has been raised by the infusion of KCl, it falls approximately exponentially for a time and then tends to rise. The accompanying rise and fall of calcium concentration can be fitted to a simple mathematical model. 2. The mobilization of calcium is accompanied by the generation of bicarbonate in equivalent amounts, without much change in pH or Pco2. 3. There is probably an increased production or retention of carbon dioxide at this time, but the main cause of the changes in calcium and bicarbonate is not respiratory acidosis. 4. The concentration of potassium rises in snalis that have been infused with CaCl2. 5. The adverse responses visible in snails infused with KCl are largely prevented when CaCl2 is infused at the same time. 6. Homeostasis seems to involve the maintenance of a proper balance of calcium and potassium concentrations.


A study of the unidirectional fluxes of Na and Cl across the gills of the dogfish Scyliorhinus canicula (Chondrichthyes). The gills of the dogfish Scyliorhinus canicula are more permeable to Cl than to Na. In sea water, influx of Na and Cl exceeded the efflux of these ions. Under these conditions the fish were slightly electronegative, by about 2 mV, to the external solution. The net accumulation of Cl could be accounted for by diffusion along the observed electrochemical gradient byt the movement of Na into the fish was more consistent with an electrically neutral active Na transport mechanism (using the Ussing flux ratio criterion). When the external pH was was changed from 7-8 to 6-9,, influxes of Na and Cl were depressed, while the effluxes were unaffected, and the fish became slightly less electronegative. In artificial solutions, in which the concentrations of Na and Cl were lowered and replaced with urea to maintain the total osmotic concentration, Na influx displayed saturation kinetics, while Na efflux increased with decreasing Na concentrations. Cl influx decreased linearly, while Cl efflux remained constant. The efflux of Cl could not be reconciled with a process of passive diffusion along any of the observed electrochemical gradients and thus could reflect the presence of an active transport mechanism.


The actions of some putative neurotransmitters on the cockroach salivary gland. 1. Certain putative transmitters were applied to the innervated cockroach salivary gland and their effects on the resting potential and the neurally evoked secretory potential of the acinar cells were observed. 2. gamma-Aminobutyric acid, glutamate, glycine, aspartate and alanine had no significant effect on the resting potential. However, gamma-aminobutyric acid and glutamate reduced the neurally evoked secretory potential but only at concentrations above 10(-3) M3. Acetylcholine and carbachol appeared to act by modifying transmitter output from the salivary nerves. These substances failed to have any effect on the resting potential. 4. The biogenic amines, adrenaline, dopamine, noradrenaline, 5-hydroxy-tryptamine and octopamine, produced hyperpolarizing responses, graded according to concentration. 5. It is suggested that dopamine, the most potent of the biogenic amines tested, is the transmitter at this junction.


Branchial ion uptake in arctic grayling: resting values and effects of acid-base disturbance. 1. Techniques for the measurement of unidirectional flux rates in fish which require no anaesthesia or surgery are described. 2. Resting values for Cl- uptake at 10 and 17 degrees C were 8-03 +/- 1-11 and 13-52 +/- 0-95 mu-equiv. 200 g-1 h-1 (+/- S.E.), respectively; and for Na+ the rates were 15-49 +/- 0-40 and 26-30 +/- 0-36, respectively. 3. Hypercapnic acidosis caused an increase in Na+ uptake, presumably through Na+/H+ (or NH+4) exchange. It is suggested that this is a compensation mechanism leading to the increase in blood buffering observed in response to hypercapnia. 4. Alkalosis was observed following acute temperature increase and was accompanied by an increase in the rate of Cl-/HCO-3 exchange and also by an increase in Na+/H+ exchange. 5. The role of these branchial ion exchange mechanisms in overall acidbase regulation is discussed.


[Characterization of the oxidoreductase inhibitor (author's transl)]. The oxidoreductase inhibitor was prepared from NAD in alkaline solution, and purified chromatographically. Investigations are reported on the extinction and fluorescence spectra, stability of the inhibitor, and the dependence of inhibition on the concentration of enzyme and coenzyme. Kinetic studies show that the inhibition is non-competitive.


[Formation and purification of the oxidoreductase inhibitor from NAD (AUTHOR'S TRANSL)]. The oxidoreductase inhibitor is not formed from NADH as previously thought, but only from NAD under alkaline conditions. Analogues of NAD (e.g. NADP) and components of the NAD molecule (e.g. ADP) have no effect on the formation of the inhibitor. The most favourable pH, temperature, duration of incubation, type of buffer and NAD concentration for the formation of the inhibitor were investigated. The method for the formation and chromatographic isolation of the oxidoreductase inhibitor is briefly described.


Ventilation and metabolic rate of young rainbow trout (Salmo gairdneri) exposed to sublethal environmental pH. Differences between ventilatory response and metabolic rates of young rainbow trout tested within the sublethal range of pH 6 to pH 9 were observed using a flowing water respirometer. The oxygen consumption was monitored at swimming speeds of 12 cm/sec and 24 cm/sec. The oxygen consumption rates at 24 cm/sec and pH 6 (423 mg/kg-hr) and pH 9 (367 mg/kg-hr) were considerably higher than those determined near neutrality (328 mg/kg-hr). Ventilation rate increased to either side of neutrality, but significantly fewer respiratory reversals, or "coughs," were observed at pH 6 and a greater number at pH 9 than occurred at pH 7 and 8 or in untested fish. The respiratory-cough response is shown to be pH-dependent in rainbow trout and may therefore not be as reliable an indication of pollutant-caused stress in studies where the experimental pH has not been specified or controlled.


Gustatory hairs on the mosquito, Culiseta inornata. Gustatory hairs were investigated on the legs and mouthparts of Culiseta inornata (Williston) (Diptera: Culicidae). One type of hair, each innervated by four neurons, was found on the legs. Two of the neurons responded to NaCl stimulation, one neuron to water stimulation, and one neuron to sucrose stimulation. Three kinds of hairs designated Type I (T1), Type 2 (T2) and Type 3 (T3) were analyzed on the labella. The T1 hairs are innervated by one sugar neuron, one mechanoreceptor, two salt neurons and one water neuron. The T2 hairs are innervated by two salt neurons and one mechanoreceptor. The T3 hairs, located on the oral surface of the labella, are innervated by a variable number (2-5) of neurons. Precise identification of the T3 chemosensory neurons was not made because of the small size and inaccessibility of the T3 hairs. Chemosensory hairs on the tip of the labrum were tested electrophysiologically. the sequence of decreasing effeectiveness for the three salts tested was KCl greater than NaCl greater than LiCl. Labral chemoreceptors also responded positively to sucrose.


Proton transport by phosphate diffusion--a mechanism of facilitated CO2 transfer. We have measured CO2 fluxes across phosphate solutions at different carbonic anhydrase concentrations, bicarbonate concentration gradients, phosphate concentrations, and mobilities. Temperature was 22-25 degrees C, the pH of the phosphate solutions was 7.0-7.3. We found that under physiological conditions of pH and pCO2 a facilitated diffusion of CO2 occurs in addition to free diffusion when (a) sufficient carbonic anhydrase is present, and (b) a concentration gradient of HCO3- is established along with a pCO2 gradient, and (c) the phosphate buffer has a mobility comparable to that of bicarbonate. When the phosphate was immobilized by attaching 0.25-mm-long cellulose particles, no facilitation of CO2 diffusion was detectable. A mechanism of facilitated CO2 diffusion in phosphate solutions analogous to that in albumin solutions was proposed on the basis of these findings: bicarbonate diffusion together with a facilitated proton transport by phosphate diffusion. A mathematical model of this mechanism was formulated. The CO2 fluxed predicted by the model agree quantitatively with the experimentally determined fluxes. It is concluded that a highly effective proton transport mechanism acts in solutions of mobile phosphate buffers. By this mechanism; CO2 transfer may be increased up to fivefold and proton transfer may be increased to 10,000-fold.


Distribution of rhodopsin and retinochrome in the squid retina. The cephalopod retina contains two kinds of photopigments, rhodopsin and retinochrome. For many years retinochrome has been thought to be localized in the inner segments of the visual cells, whereas rhodopsin is in the outer segments. However, it is now clear that retinochrome can be extracted also from fragments of outer segments. In the dark-adapted retina of Loligo pealei retinochrome is distributed half-and-half in the inner and outer segments. Todarodes pacificus contains much more retinochrome than Loligo, and it is more abundant in the outer than in the inner segments. The outer segments of Loligo contain retinochrome and metarhodopsin in addition to rhodopsin, whether squids are kept in the dark or in the light. But there is extremely little metarhodopsin (about 3% of rhodopsin) even in light-adapted eyes. The inner segments contain only retinochrome, and much less in the light than in the dark. On the other hand, retinochrome in the outer segments increases markedly during light adaptation. These facts suggest the possibility that some retinochrome moves forward from the inner to the outer segments during light adaptation and there reacts with metarhodopsin to promote regeneration of rhodopsin.


Formation and isolation of leucocidin from Pseudomonas aeruginosa. A toxic substance, which destroyed leucocytes from man but was inactive against erythrocytes, was demonstrated in cultures of four out of 110 strains of Pseudomonas aeruginosa tested. The toxin, designated 'leucocidin', was cell-bound as a precursor toxin, exhibiting little or no toxicity. It was converted into toxin with maximum activity by various proteases including an endogenous elastase. The production of leucocidin was directly proportional to the number of bacteria and was not influenced by variations in media, iron concentration, pH or temperature. The best method for large-scale production of leucocidin was autolysis of washed bacteria.


The production and growth characteristics of yeast and mycelial forms of Candida albicans in continuous culture. The growth characteristics of Candida albicans CM145,348 have been examined under aerobic conditions in continuous culture. At different steady states the environment was controlled with respect to the concentrations of dissolved oxygen, carbon and nitrogen, the pH, and the temperature. Dry matter, substrate concentration, yield, specific oxygen uptake, specific carbon dioxide release and respiration quotient were examined as a function of the dilution rate. The morphology depended on the carbon source. Maltose produced a mycelial morphology, whereas with lactate a yeast culture was obtained. With fructose or glucose as a carbon source a mixed morphology of yeast, pseudo-mycelial and mycelial forms was produced. A larger number of different growth conditions were examined in batch culture but a mixed morphology was always obtained.


The effect of nitrogen limitation on catabolite repression of amidase, histidase and urocanase in Pseudomonas aeruginosa. In Pseudomonas aeruginosa, the synthesis of histidase, urocanase and amidase is severly repressed when succinate is added to a culture growing in pyruvate + ammonium salts medium. When growth is nitrogen-limited, catabolite repression by succinate of histidase and urocanase synthesis does not occur but succinate repression of amidase synthesis persists. Amidase synthesis is not regulated in the same way as histidase synthesis by the availability of other nitrogen compounds for growth. Growth of P. aeruginosa strain PACI in succinate + histidine media is nitrogen-limited since this strain is defective in a histidine transport system. When methyl-ammonium chloride is added to succinate + histidine media, growth inhibition occurs. Mutants isolated from succinate + histidine + methylammonium chloride plates were found to be resistant to catabolite repression by succinate even in ammonium salts media. It is suggested that the hut genes of P. aeruginosa may be regulated in the same way as in Klebsiella aerogenes, by induction by urocanate and activation by either the cyclic AMP-dependent activator protein or by glutamine synthetase.


Decreased permeability as a mechanism of resistance to methyl benzimadazol-2-yl carbamate (MBC) in Sporobolomyces roseus. Mutants of Sporobolomyces roseus resistant to benzimidazole fungicides varied in their responses to 2-(thiazol-4-yl) benzimidazole (thiabendazole, TBZ), methyl 1-(butylcarbamoyl)-benzimidazol-2-yl carbamate (benomyl) and methyl benzimidazol-2-yl carbamate (carbendazim, MCB). Incorporation of [14C] MBC into trichloroacetic acid extracts of the sensitive strain S4 increased during a 2 h incubation period, whereas incorporation into the resistant mutant M55 was unchanged. [14C] MBC uptake by S4 cells was five times higher than that by M55. MBC was identified as the main radioactive compound inside the S4 cells and reached a level of 2.4 mug/100 mg dry wt. The compound MBC enters the cells of Sp. roseus by a temperature-, energy-, pH- and concentration-dependent transport system which may be specific for compounds containing a benzimidazole nucleus. It is suggested that tolerance of M55 to MBC is due to decreased permeability of the cell to this compound.


Accumulation and storage of Zn2+ by Candida utilis. Starved cells of Candida utilis accumulated Zn2+ by two different processes. The first was a rapid, energy- and temperature-independent system that probably represented binding to the cell surface. The cells also possessed an energy-, pH-, and temperature-dependent system that was capable of accumulating much greater quantities of the cation than the binding process. The energy-dependent system was inhibited by KCN, Na2HAsO4, m-chlorophenyl carbonylcyanide hydrazone, N-ethylmaleimide, EDTA and diethylenetriaminepenta-acetic acid. The system was specific inasmuch as Ca2+, Cr3+, Mn2+, Co2+ or Cu2+ did not compete with, inhibit, or enhance the process, Zn2+ uptake was inhibited by Cd2+. The system exhibited saturation kinetics with a half-saturation value of 1.3 muM and a maximum rate of 0.21 (nmol Zn2+) min(-1) (mg dry wt(-1)) at 30 degrees C. Zn2+ uptake required intact membranes since only the binding process was observed in the presence of nystatin, toluene, or sodium dodecyl sulphate. Cells did not exchange recently accumulated toluene, or sodium dodecyl sulphate. Cells did not exchange recently accumulated 65Zn following the addition of a large excess of non-radioactive Zn2+. Similarly, cells pre-loaded with 65Zn did not lose the cation during starvation, and efflux did not occur when glucose and exogenous Zn2+ were supplied after the starvation period. Efflux was only observed after the addition of toluene or nystatin, or when cells were heated to 100 degrees C. Cells fed a large quantity of Zn2+ contained a protein fraction resembling animal cell metallothionein. In batch culture, cells of C. utilis accumulated Zn2+ only during the lag phase and the latter half of the exponential-growth phase.


Mortality and cerebral metabolism after bilateral carotid artery ligation in normotensive and spontaneously hypertensive rats. Mortality and cerebral glycolytic metabolism were studied after bilateral ligation of the common carotid artery in normotensive Wistar rats (NTR), and spontaneously hypertensive rats (SHR) derived from Wistar strain. In the first 24 hours after occlusion of carotid arteries, 72 per cent of 108 SHR died, whereas it was fatal in only 16 per cent of 43 NTR. In SHR, cerebral lactate and cerebral lactate/pyruvate ratio (L/P ratio) increased by 12.4 and 12.1 times the control, respectively at five to six hours after ligation, and remained raised even in rats surviving for two to three days thereafter. Changes in cerebral lactate and L/P ratio were minimal in NTR. Cerebral ATP decreased markedly at five to six hours after ligation in SHR studied. These results indicate that bilateral carotid artery ligation causes severe brain damage in SHR but not in NTR, suggesting hypertension per se to be operative for the development of cerebral ischaemia.


Effects of two desensitization techniques, biofeedback and relaxation, on intractable epilepsy: follow-up study. Two techniques of desedsitization, biofeedback and relaxation, were employed in a crossover design for the treatment of three young female patinets suffering from drug resistant epilepsy associated with anxiety and phobic symptoms. The patients were followed up for 15 months after the six months of treatment. The results indicate that both relaxation and biofeedback improved the patients' control of their seizures and the effects were maintained during the follow-up period.


Skeletal muscle necrosis following membrane-active drugs plus serotonin. Administration of imipramine plus serotonin (5-HT) to rats has been proposed as an animal model of Duchenne muscular dystrophy. We studied the skeletal muscle necrosis produced in male rats given 5-HT after pretreatment with imipramine, other tricyclic antidepressants, or antihistamines, which like the tricyclic antidepressants, can block neuronal reuptake of 5-HT. Following one of these agents plus 5-HT, 20 mg/kg subcutaneously (s.c.), necrosis was more severe in the soleus muscle than the quadriceps. There was no significant difference in the incidence of necrosis in the soleus and quadriceps muscles following one of these agents plus 5-HT, 100 mg/kg, intraperitoneally (i.p.). After one of these agents plus 5-HT i.p., but not 5-HT s.c., extensive necrosis was significantly more frequent and severe in the quadriceps muscle than after 5-HT s.c. Chlorpheniramine (CP) plus 5-HT, 2.5 mg/kg intravenously, produced less muscle necrosis than CP plus 5-HT s.c. or i.p. The necrosis produced by CP plus 5-HT s.c. was comparable ipsilateral and contralateral to the injection site. The necrosis following CP plus 5-HT i.p. was maximal at 24 hr and remained fairly constant until 5 days. Regeneration was prominent by 7 days. The muscle necrosis  produced by CP plus 5-HT is blocked by some 5-HT blockers, e.g., methiotepin and methysergide. It is also partially blocked by denervation. The capacity of tricyclic antidepressants and antihistamines to block neuronal 5-HT reuptake tended to be negatively correlated with the capacity to potentiate the muscle necrosis they produced with 5-HT, which suggests that blockade of 5-HT uptake is not the mechanism of the pathology produced by the combined treatment. The tricyclic antidepressants and the antihistamines are "membrane stabilizers-labilizers". Other drugs which are "membrane stabilizers-labilizers" such as trihexyphenidyl and procaine also promoted skeletal muscle necrosis when given prior to 5-HT. It is proposed that the effects of imipramine plus 5-HT on skeletal muscle are not due to the blockade of neuronal uptake of 5-HT and subsequent vascular-induced ischemia, but reflect direct toxic effects of these agents on skeletal muscle.


Time sequence of lipogenic changes in adipose tissue of rats when converted from ad libitum feeding to meal-eating. This study was undertaken to establish the time sequence of lipogenic changes in adipose tissue of rats when converted from ad libitum feeding to meal-eating. Rats were fed a high carbohydrate diet 2 hours/day for 0 to 10 days (meal-eating). The high speed supernatant fraction from homogenized epididymal fat pads was assayed for citrate cleavage enzyme, acetyl CoA carboxylase, fatty acid synthetase and malic enzyme activities. The effects of meal-feeding on in vitro and in vivo rates of fatty acid synthesis in adipose tissue as well as the amounts of glycogen deposited in the adipose tissue were measured. During the first 10 days of meal-feeding, the lipogenic enzyme activities were actually decreased or unchanged in the meal-fed rats but during this time the in vitro and in vivo rates of fatty acid synthesis were progressively increased in the meal-fed rats. Glycogen levels in the adipose tissue of meal-fed rats were greater than the levels in the nibblers. The initial hyperlipogenesis observed in the meal-fed rat appears to be due to changes in substrate uptake by the adipose tissue and/or to alterations in enzyme activation in the adipose tissue rather than to changes in the quantity of enzyme present in the tissue.


Guanosine nucleotide precursor for flavinogenesis of Eremothecium Ashbyii. The purine precursor in the riboflavin biosynthetic pathway in Eremothecium ashbyii was examined using a guanine analogue, 8-azaguanine, with non-growing cell systems. 1. Riboflavin formation in the culture filtrate was determined at 0, 5, 10 and 20 hr after start of the incubation of the non-growing cells in the presence of xanthine or 8-azaguanine (1 mM, respectively). At 20 hr of incubation, the addition of xanthine stimulated riboflavin formation by 36% and the addition of 8-azaguanine inhibited the formation by 57%. 2. Acid soluble nucleotide pools in the cells were followed at 0, 5, 10 and 20 hr of the incubation period in the presence of xanthine or 8-azaguanine by means of anion exchange column chromatography. The result showed that the GTP pool changed markedly despite the fact that the adenosine nucleotide pool was almost constant irrespective of the presence or absence of these purines till 10 hr of incubation. But, the decrease of the former was overcome in part by the addition of flavinogenic xanthine. Furthermore, the total amounts of GTP and guanosine accumulated in cells in the presence of 8-azaguanine reached the maximum already at 5 hr, attaining a level twice as much as the GTP contents of the control. 3. The role of guanosine nucleotide pool in riboflavin formation was further examined using 8-azaguanine. In this experiment the drug was added to the suspension of non-growing cells at 3 hr or 6 hr after the incubation was started and the reaction was continued till the 12th hr. A more clear-cut correlationship between riboflavin formation and guanosine nucleotide pool was oberved by this experiment. The guanosine nucleotide pool (consisting of GMP, GDP and GTP) increased simultaneously with the inhibition of riboflavin formation. Of the guanosine nucleotides pools, the GMP pool increased 2.7 times above normal upon the addition of 8-azaguanine during the incubation for 6 hr and 5.3 fold for 9 hr. While, the GTP pool increased 1.9 fold above normal for 6 hrs' incubation in the supplementation of this drug but decreased to one-half of the normal at the incubation period of 9 hr. In these cases, the decreased amounts of GTP were equal to the increased amounts of GMP during the incubation periods of 6 hr and 9 hr in the presence of added 8-azaguanine. 4. The above results suggest strongly that GTP is an immediate precursor of riboflavin in the form of nucleotide.


Lysozyme damage caused by secondary degradation products during the autoxidation process of linoleic acid. Autoxidized LA is classified into four groups, LA, LAHPO, SP and FP. Lysozyme is inactivated by these products in the increasing order as follows: FP less than LA less than LAHPO less than SP. The effects of these products on the amino acid composition of lysozyme is examined. All kinds of amino acid residues were not damaged until lysozyme was incubated with LA and LAHPO at 45 degrees C for 100 days. The susceptible amino acid residues attacked by the autoxidized products are tryptophan, lysine and histidine. The specific loss of methionine by SP occurs during acid-hydrolysis. The effect of SP was the strongest among the autoxidized products. FP was almost noneffective. The destructive actions of BP, MA and PA were compared with those of autoxidized products. Effects of these compounds did not resemble those of autoxidized products. It was concluded that tryptophan, lysine and histidine residues were specifically attacked by SP.


Hypomagnesemia in infants of diabetic mothers: perinatal studies. Fifty-six diabetic mothers and their infants were studied prospectively from birth. Twenty-one of 56 IDM had serum Mg less than or equal to 1.5 mg/dl, on at least one occasion during the first 3 days. Serum Mg in these hypomagnesemic infants did not demonstrate the normal increase with postnatal age that was present in normomagnesemic infants. Decreased neonatal serum Mg was related to increased severity of maternal diabetes, young mothers, mothers for lower gravidity, and prematurity. Decreased serum Mg, alone or with decreased ionized or total Ca, did not correlate with neuromuscular irritability in the infants. Decreased serum Mg in IDM was associated with decreased maternal serum Mg, decreased neonatal ionized and total Ca, increased serum P, and decreased parathyroid function. Serum Mg was not related to dietary P intake, or urinary Ca or P excretion. Thus, transitory neonatal hypomagnesemia occurs in IDM; it is speculated that factors causing HM might include maternal HM or neonatal hyperphosphatemia, and that the HM is related to the hypocalcemia and functional hypoparathyroidism of IDM.


The stability of cannabis and its preparations on storage. Solutions of pure cannabinoids, nine samples of herbal and two of resin cannabis (one freshly prepared) were stored in varying conditions for up to 2 years. Exposure to light (not direct sunlight) was shown to be the greatest single factos in loss of cannabinoids especially in solutions, which should therefore be protected from light during analytical and phytochemical operations. Previous claims that solutions in ethanol were stable have not been substantiated. The effect of temperature, up to 20 degrees, was insignificant but air oxidation did lead to significant losses. These could be reduced if care was taken to minimize damage to the glands which act as "well filled, well closed containers". Loss of tetrahydrocannabinol after exposure to light does not lead to an increase in cannabinol, but air oxidation in the dark does. It is concluded that carefully prepared herbal or resin cannabis or extracts are reasonably stable for 1 to 2 years if stored in the dark at room temperature.


Changes in the particle size distribution during tableting of sulphathiazole powder. Five size fractions of sulphathiazole powder (volume surface mean diameter 155, 133, 86, 50 and 41 mum) were compressed into 12 mm diameter tablets on an instrumented single punch tablet machine. The size analysis of the tablet material after compression showed an attrition of the coarser fraction and an agglomeration of the finer fraction. It is postulated that there is a critical particle size where the effects of crushing and bonding cancel each other. The changes in particle size are discussed in relation to some of the compressive characteristics of the powder.


Release of a drug from homogeneous ointments containing the drug in solution. The rate of release of resorcinol (5%) from hydrogels (Carbopol, sodium carboxymethylcellulose, starch), lipogels (alcoholic-base, esteric-bases containing different amounts of beeswax with and without a spreading additive, respectively) and Labrafils has been examined. For the experimental design adopted the release of the drug is linear between 10 and 70% of the amount of drug released. The results agree well with the mathematical model postulated by Higuchi (1962) for the release of a drug from homogeneous ointments containing the drug in solution.


The persistence of dextran 70 in blood plasma following its infusion, during surgery, for prophylaxis against thromboembolism. An infusion of dextran (mean molecular weight 70000) in normal saline (either 1 litre or 500 ml) was given to patients undergoing hysterectomy. The infusion was started at induction of anaesthesia and continued throughout the operation and for up to 5 h thereafter. The rate of elimination of dextran was independent of the dose given. The time to eliminate half the dose was nearly two days and up to 10% was still present in the circulation after one week. The persistence of dextran in the plasma in these amounts and for this length of time may have considerable implications in the prophylaxis of postoperative deep venous thrombosis.


The absorption and elimination of metoclopramide in three animal species. The absorption and elimination of metoclopramide have been studied in the rat, rabbit and dog. Thin-layer chromatography followed by photodensitometry was used for the analysis of the unchanged drug and its metabolites. N-De-ethylation is an important Phase I metabolic reaction and conjugation with glucoronic acid and sulphate is a major route of metabolism, particularly in the rabbit. The pharmacokinetic parameters after intravenous administration showed little interspecies variation. First order elimination kinetics with short half-lives and high apparent volumes of distribution (greater than 1.1 kg(-1)) were observed. Major interspecies variations were seen after oral administration of high doses of the drug. Metoclopramide was eliminated slowly after oral administration to rats. The findings in the rabbit and in the dog suggest that the liver plays an active role reducing the systemic availability of unchanged metoclopramide after oral administration.


Release of prostaglandin E-like material from perfused mesenteric blood vessels of rabbits. Infusions of noradrenaline (1-3 mug ml(-1) min(-1)) into the mesenteric vascular preparation of the rabbit caused a 2 to 5 fold rise in perfusion pressure and a release of prostaglandin E-like material (3.23 +/- 0.65 (s.e.) ng PGE2 equivalents ml(-1)). Indomethacin (3 mug ml(-1)) prevented whereas arachidonic acid (0.2 mug ml(-1)) augmented, the noradrenaline-evoked release of a prostaglandin E-like material. The walls of arterioles or precapillary vessels are the proposed site of prostaglandin generation.


Kininogen and kininogenase synthesis by the liver of normal and injured rats. Isolated livers from normal rats or from others at 2 days after subcutaneous injection of turpentine have been perfused with a simplified medium. Estimation of kininogen, kininogenase and 2 plasma proteins in the perfusates thus obtained indicate that both kininogen and kininogenase are synthesized in the liver. Furthermore, because twice as much kininogen and kininogenase was synthesized by the livers from the rats which had been injected with turpentine as by those from the normal rats, these two proteins must be considered to be members of the group of plasma proteins known as acute phase reactants.


The effect of some polyene macrolides on absorption from the small intestine in the rat. The effect of three polyene macrolides, candicidin, amphotericin B and nystatin on the absorption of [3H] cholesterol was studied in the rat by using the in situ gut loop perfusion technique. Chronic treatment with candicidin and its presence at various concentrations in the gut loop perfusion experiments inhibited [3H] cholesterol absorption although a smaller effect was also obtained with amphotericin B and nystatin at higher concentrations. A similar but much less pronounced action of candicidin was also observed on the absorption of [3H] corticosterone and [14C] phenytoin.


An assessment of the cardiovascular sympathectomy induced by guanethidine. Guanethidine treatment of rate (30 mg kg(-1), i.p. daily for 6 weeks) produced a profound reduction in the catecholamine present (as indicated by fluorescence histochemistry and catecholamine determinations) in tissues taken from the cardiovascular system, but there was evidence of the return of catecholamines within 8 weeks. While these changes are consistent with a sympathectomy, the unaltered pressor responses to physostigmine (100 mug kg(-1), i.v.) and to carotid occlusion indicate an unimpaired functional capacity of noradrenergic nerves supplying the cardiovascular system. Although part of the response may be attributed to the unaffected adrenal medulla enhanced by the presence of considerable supersensitivity as shown to exogenous noradrenaline, there would appear to be a dissociation between the results obtained from physical and functional tests of the sympathectomy induced by guanethidine.


Some formulation factors affecting the tensile strength, disintegration and dissolution of uncoated oxytetracycline tablets. A study has been made of the effects of gelatin binding agent and moisture content on the tensile strength, disintegration and dissolution times of oxytetracycline tablets. These properties all increase with the gelatin content, and maximum tensile strengths occur when the tablets contain between 2.5 and 4.5% w/w of moisture. The properties of the tablets depend on their packing fraction and in general, the disintegration and dissolution times are minimal when the packing fraction is between 0.77 and 0.82. A connection has been established between the disintegration times of the tablets and the time required for 50% of the drug content to dissolve.


Factors affecting the binding of tricyclic tranquillizers and antidepressants to human serum albumin. The linear free energy-related model for structure activity relations developed by Hansch & Fujita (1964) has been used to correlate the binding of tricylic tranquillizers and antidepressants to human serum albumin (HSA) with hydrophobic and electronic parameters. The parameters chosen being the chromatographic parameter (Rm) and the affinity of charge transfer complex formation (kc). The relative importance of these factors has been assessed by linear and multiple linear regression analysis. Results show that the major factor in binding is electronic with only a minor contribution from the hydrophobic parameter.


Reversed ester analogues of pethidine: isomeric 4-acetoxy-1, 2, 6-trimethyl-4-phenylpiperidines. The preparation and stereochemical characterization of all three isomeric forms of 4-acetoxy-1, 2, 6-trimethyl-4-phenylpiperidine is described. Of these, only the t-2-Me, c-6-Me, r-4-OCOMe isomer was an effective analgesic in mice (2.3 X pethidine) as judged by the hot-plate test. The results, together with reported data, demonstrate the potency raising effects of axial methyl alpha- to nitrogen and the lowering action of equatorial alpha-methyl substituents in reversed esters of pethidine.


Studies with the International Pyrogen Standard on the sensitivity and reproducibility of pharmacopoeial pyrogen testing. Rabbits, 27 or 36 in each experiment, were injected with the International Pyrogen Standard (I.P.St.) in different seasons. The maximum temperature rises were registered, randomized and interpreted according to the requirements of the B.P. (1973), U.S.P. (1970), P. Hung. (1970) and P. Nord. (1962). Although the dose of 3.5 ng kg(-1) I.P.St. proved to be non-pyrogenic as tested in summer, when tested in winter the same dose was qualified pyrogenic (to be rejected) by up to one third of the combinations by the criteria of the four Pharmacopoeias. In the spring experiment "to be rejected" qualifications predominated as based on the response of large groups of rabbits. Exclusion of the rabbits showing low sensitivity (before randomization) barely influenced the results with 3.5 ng kg(-1) I.P.St. in the experiment in which the mean temperature rise was 0.49 degrees. If, however, the mean temperature rise was higher (0.57 or 0.69 degrees), such a selection practically resulted in the disappearance of "passable" qualifications in the triplet groups and a great predominance of "to be rejected" qualifications in the larger groups. The dose 7.0 ng kg(-1) consistently proved to be pyrogenic in large groups of rabbits.


Gastrointestinal absorption of carbenoxolone in the rat determined in vitro and in situ: deviations from the pH-partition hypothesis. The absorption of [14C] carbenoxolone from everted rat ileum in vitro and from rat stomach and ileum in situ has been examined. The rate of its mucosal to serosal transfer in vitro increases as pH increases from 5 to 8 whereas the amount bound to ileum tissue decreases with increased pH; absorption closely parallels the drug's solubility. The uptake of carbenoxolone in situ is bi-exponential and the rate constants for the two processes, have been calculated. Absorption in situ, and biliary excretion, of the drug increases with increasing pH from 5.0 to 7.4. Tissue binding to the ileum in situ is not dependent on pH except below pH 5.0 when extensive tissue accumulation of carbenoxolone occurs because of its low solubility. Tissue binding to the stomach increases markedly with decrease of pH from 7.4 to 6.5 and at pH 6.5 is 80 times greater than binding to the intestine. The rate of absorption from the stomach, at pH 6.5-7.4, was much less than that from the intestine in situ. When allowance is made for the binding of carbenoxolone to the stomach, contrary to the pH-partition hypothesis, correlation is apparent between its absorption and the amount present in the ionized form.


Effects of phosvitin on the ecg changes induced under hypoxia in the rat. The effect of phosvitin (1 g kg(-1), i.p.) on ecg changes induced in rats by a reduction of partial oxygen pressure in the respiratory mixture was studied. Phosphocreatine, phosphoserine, ATP and Na2HPO42H2O were also administered intraperitoneally for comparison. Phosvitin alone was found to prevent the hypoxia-induced T-wave changes (flattening or disappearance), which were also temporarily aggravated by injection of noradrenaline. As to the metabolic, hypoxia-induced myocardial changes, two hypotheses are discussed: a release of phosvitin phosphate radicals ready for immediate utilization or a drug action mediated via a membrane-bound intrinsic proteinkinase system.


The transport of tetracyclines across the mouse ileum in vitro: the effect of cations and other agents. The intestinal transfer of different tetracyclines dissolved in calcium- and magnesium-free Krebs bicarbonate buffer solution, pH 7.4, was studied using the everted ileum of the mouse. The rates of transfer of chlortetracycline and demethylchlortetracycline were less than those of tetracycline and oxytetracycline, the latter compounds being transferred at the same rate. Addition of calcium and magnesium to the buffer greatly reduced the transfer of tetracycline; this inhibition could be antagonized by EDTA. The presence of iron also inhibited the transfer of tetracycline. The inhibitory effect of these ions on tetracycline transfer seemed due to chelation of the drug. Glucosamine and acetylmethionine, but not acetyl glucosamine, diminished the intestinal transfer of tetracyclines. The former two agents did not influence the uptake of tissue fluids. Tetracycline was also transfered from the serous to the mucous coat in the non-everted intestinal sac of mice. The above observations suggested that the absorption of tetracyclines was not due solely to passive diffusion.


Some pharmacological properties of the venom, venom fractions and pure toxin of the yellow-bellied sea snake Pelamis platurus. The effects of the crude venom, four partially purified venom fractions and pure toxin (Pelamis toxin alpha) from yellow-bellied sea snake, Pelamis platurus, on respiration, blood pressure, heart and skeletal muscle of rabbits have been examined. Results indicated that crude venom, a partially purified toxic fraction and Pelamis toxin alpha caused initial respiratory stimulant effects followed by respiratory paralysis. In most cases, respiratory paralysis occurred before a profound fall in arterial pressure. Depression of the twitch response to nerve stimulation was observed in the tibialis anterior muscle. No significant change in the electrocardiogram was seen. Three partially purified non-toxic fractions of the crude venom induced transient respiratory stimulant effects. It was concluded that the crude venom and Pelamis toxin alpha had an identical mode of action and that they caused respiratory paralysis in rabbits.


The influence of crystal form on the radial stress transmission characteristics of pharmaceutical materials. Various crystal forms of sulphathiazole, barbitone and aspirin were compressed in a single-punch tablet machine instrumented to monitor axially applied and radially transmitted forces, and upper punch movement. The changes in radial stress during the compression cycle depended upon the polymorphic form of the compressed material. The results were rationalized in terms of the degree of plastic flow/crushing that occurred with each material, and the degree to which the final compact underwent elastic compression. It is postulated that the reduction in the transition temperature of polymorphic forms of sulphathiazole and barbitone and the polymorphic transition of sulphathiazole Form II was due to the production of dislocations in the crystal and the crystals at crystal boundaries formed in the compressed materials.


Use of the mouse jumping test for estimating antagonistic potencies of morphine antagonists. The potencies of 19 reference morphine antagonists have been compared in a modified version of the mouse jumping test. Mice were each implanted subcutaneously with one 75 mg pellet of morphine. Antagonist challenge took place 72 h later and the incidence of repetitive vertical-jumping was monitored over 1 h. A high Pearson correlation coefficient (r = 0.997) was found between quantitative assays based on the total number of jumps per mouse and quantal assays based on mice jumping at least 6 times. A comparison of relative potencies obtained with the mouse test and with non-withdrawn morphine-dependent monkeys gave a Spearman rank order coefficient of 0.91 while a similar comparison with values obtained with the guinea-pig isolated ileum preparation also gave a high correlation coefficient (r= 0.92). Whereas it is difficult to assess the antagonistic component of buprenorphine and cyclorphan with the ileum preparation, both compounds can be satisfactorily assayed in the mouse jumping test. The reported antagonistic properties of ketocyclazocine and profadol could not be confirmed in the mouse model.


The narcotic discriminative stimulus complex: relation to analgesic activity. The ability of drugs to produce the narcotic discriminative stimulus complex is found to be highly correlated with their analgesic activity; in contrast, no relation with their antidiarrhoeal activity is evident. The findings suggest that the narcotic discriminative stimulus complex is a centrally mediated effect of narcotic drugs.


A sensitive biological assay for prostaglandin E and acetylcholine. Superfused hamster stomach strip has been used in the bioassay of prostaglandin E2 (PGE2) and acetylcholine. The preparation proved very stable and had no spontaneous movements. The sensitivity to PGE2 (and PGE1) was in the range of 10(-9) g ml(-1) and to acetylcholine 10(-12) g ml(-1). After treatment with physostigmine the sensitivity to acetylcholine was 10(-15) g ml(-1). In acetylcholine determination the preparation is more sensitive than any other method.


Intestinal pH and propulsion: an explanation of diarrhoea in lactase deficiency and laxation by lactulose. Subjects deficient in lactase may experience bloating, cramps and diarrhoea after ingesting milk, due to the unhydrolysed and poorly-absorbed lactose. The diarrhoea may result from an osmotic effect of the lactose itself or its poorly-absorbed acidic products of fermentation (Weijers, van de Kamer & others, 1961; Christopher & Bayless, 1971), possibly together with an alteration of sodium and water absorption due to the lowered colonic pH (Rousseau & Sladen, 1971). Laxation by lactulose (1-4-beta-galactosidofructose) may operate through an analogous mechanism. The drug is a synthetic dissaccharide which, in oral doses of 10-20 g, relieves chronic constipation (Wesselius-de Casparis, Braadbaart & others, 1968). It is neither hydrolysed by intestinal dissaccharidase (Dahlqvist & Gryboski, 1965) nor absorbed in the gut, but it is converted in the colon mainly to lactic and acetic acids by various bacteria including Lactobacillus acidophilus. Apart from the increased osmotic effect, the pH in the proximal colon falls markedly (Bown, Gibson & others, 1974), and larger doses may reduce stool pH. Weijers & others (1961) inferred that the acidic products formed from lactose in the colon stimulate propulsion, and K.S. Liem (Philips-Duphar) suggested to us that lactulose may relieve constipation partly by stimulation of propulsion due to the lowered pH. The experiments described below support this view.


Acetylcholinesterase and responses to acetylcholine in the embryonic chicken heart. Three and 4 day old embryonic chicken hearts were examined for their responsiveness to acetylcholine and presence of acetylcholinesterase (AChE) to determine the role of the enzyme in the cardiac effects of the transmitter. The effects of acetylcholine on rate and contractility of 3 day old hearts were indistinguishable from those on 4 day old hearts. The effects were readily blocked by atropine at both stages of development. In 3 day old hearts the responses to acetylcholine were not affected by the AChE inhibitor physostigmine but in 4 day old hearts they were considerably potentiated. The effect of acetylcholine on the rates of 4 day old hearts is of short duration (5 min or less). In 3 day old hearts it persists for a much longer time. Thus, the appearance of AChE in the embryonic heart of the chicken does not seem to modify the responsiveness of the cholinergic receptor to the transmitter.


The effect of propranolol on sympathetic nerve stimulation in isolated vasa deferentia. (+/-)-Propranolol hydrochloride (0.5 mg kg(-1) twice daily, subcutaneously, for 3 days or approximately 2.4 mg kg(-1) daily, orally, for 21 days) failed to produce ptosis or to affect responses to transmural stimulation of isolated vasa deferentia removed from treated mice. In guinea-pig isolated vasa deferentia responses to transmural stimulation through parallel electrodes were reduced by propranolol (1 to 20 mug ml(-1); blockade was concentration dependent, fast to equilibrium (45 min), easily reversed by washing but not reversed by (+)-amphetamine sulphate (0.2 mug ml(-1). At lower concentrations (0.04 and 0.2 mug ml(-1), propranolol marginally potentiated responses to transmural stimulation. In contrast, guanethidine (0.2 mug ml(-1)) produced a slow onset blockade which was completely reversed by (+)-amphetamine. The response to electrical stimulation through concentric ring electrodes was reduced by low concentrations of propranolol but this effect is ascribed to the known local anaesthetic actions of propranolol and no evidence of true adrenergic neuron blockade was found.


Some effects of urea on drug dissolution. Solubilities and dissolution rates of salicylic acid have been determined in urea solutions at different pH values. Solubilities increased with pH and urea concentration; a solubilization mechanism was considered to be operating. The solubilization effect of urea was greatest on the non-ionzed moieties of the solute. Dissolution rates of salicylic acid increased with pH and urea concentration. The increase in dissolution rate paralleled increases in solubility. The role of solubilizing effect in the enhancement of dissolution rate by urea is discussed.


Solubilization of hydrocortisone, dexamethasone, testosterone and progesterone by long-chain polyoxyethylene surfactants. Solubility and dialysis methods were used to study the solubilization of hydrocortisone, dexamethasone, testosterone and progesterone in aqueous long-chain polyoxyethylene non-ionic surfactant solutions. Partition coefficients, Km, between micellar and aqueous phases were calculated between 10-50 degrees. Km decreased with temperature and polyoxyethylene chain length but increased with decrease in steroid polarity. The standard free energy change, deltaGOS, for the solubilization of the steroids decreased with decrease in steroid polarity and surfactant hydrophilic chain length but was essentially independent of temperature. The enthalpies and entropies for the process were determined from the variation of Km with temperature. deltaHOS and deltaSOS increased with decreasing steroid polarity but were essentially independent of temperature and polyoxyethylene chain length.


Influence of non-ionic surfactants on permeation of hydrocortisone, dexamethasone, testosterone and progesterone across cellulose acetate membrane. The lag-time method of diffusion has been used to investigate permeation of hydrocortisone, dexamethasone, testosterone and progesterone across cellulose acetate membranes between 10 degrees and 40 degrees. The process depended mainly on membrane-water partition coefficients of the steroids so that the least polar compound permeated the fastest. Permeation generally increased with increasing temperature and from the temperature dependance of the diffusion coefficient, energies of activation were derived. The varied from 2.4 kcal mol(-1) for the least polar steroid, progesterone, to 7.4 kcal mol(-1) for the most polar, hydrocortisone. n-C16 Polyoxyethylene surfactants when present below and above the cmc increased the steroids permeation rates. Varying the polyoxyethylene chain length (OE equals 17-63) did not significantly affect permeation rates, suggesting that the enhancing effect of surfactants arises from their hydrophobic group.


Oxytetracyline tablet formulations: effect of variations in binder concentration and volume on granule and tablet properties. Formulation studies have been conducted on an oxytetracycline dihydrate tablet formulation containing microcrystalline cellulose and alginic acid, wet granulated with polyvinylpyrrolidone (PVP) solution. A range of granule properties including size, strength packing and porosity, and tablet properties including breaking load, porosity, disintegration and dissolution have been measured. Increased compaction pressure decreased tablet porosity. The reproducibility of the above properties was determined by tests on nine standard batches of granules and tablets. An increased concentration of PVP in the binder solution decreased the rate of tablet dissolution. Although the volume of granulating solution apparently controlled the granule size, it did not significantly alter the tablet dissolution, when the amount of PVP was constant.


Oxytetracycline tablet formulations: the influence of excipients and the method of granulation. The proportion of microcrystalline cellulose and alginic acid present as excipients in the dry mix for an oxytetracycline dihydrate tablet formulation, prepared by a conventional wet granulation process, has been shown to influence granule formation and properties. Granule size distributions have varied widely due perhaps to variation in binder distribution. Granulating with water was equally satisfactory to granulating with a PVP solution. Slugged granules produced robust tablets, which disintegrated and dissolved rapidly.


Oxytetracycline tablet formulations: the effect of wet mixing time, particle size and batch variation on granule and tablet properties. The wet mixing time has been shown to influence the properties of an oxytetracycline dihydrate tablet formulation, wet granulated with PVP solution. Increased time of wet mixing produced larger, stronger and more dense granules, which compressed into tablets with longer disintegration and dissolution times. Decreased drug particle size aggravated these trends. A decrease in drug particle size also produced larger, stronger and more dense granules. Above an oxytetracycline mean particle diameter of about 6 mum, the tablet dissolution was satisfactory. As the oxytetracycline particle size was decreased further, however, the distintegration and dissolution of the corresponding tablets was markedly slower.


Identification of monohydroxylated metabolites of cannabidiol formed by rat liver. Cannabidiol (CBD) was metabolized in vitro by rat liver enzymes. Unchanged CBD and eight monohydroxylated metabolites were isolated and positively identified. As previously reported, 7-hydroxy-CBD was the major metabolite. The second most abundant metabolite was 6alpha-hydroxy-CBD; whereas only a trace amount of 6beta-hydroxy-CBD was found. In addition hydroxylation occurred in all positions of the pentyl side chain, 4 inches-hydroxy-CBD being most abundant. 3 inches-Hydroxy-CBD was formed in half of the yield of 4 inches-hydroxy-CBD, while 1 inches-, 2 inches-, 5 inches-hydroxy-CBD were each formed in approximately one fourth of the yield of 4 inches-hydroxy-CBD.


Characterization of the butyl homologues of delta1-tetrahydrocannabinol, cannabinol and cannabidiol in samples of cannabis by combined gas chromatography and mass spectrometry. The butyl homologues of delta1-tetrahydrocannabinol, delta1-tetrahydrocannabinolic acid, cannabinol and cannabidiol have been identified in several samples of cannabis. 8 samples contained delta1-tetrahydrocannabinolic acid, one sample contained cannabinol and one sample contained both cannabinol and cannabidiol. Separation by gas chromatography and identification by gas chromotography-mass spectrometry was achieved by the preparation of trimethylsilyl, d9-trimethylsilyl, triethylsilyl and tri-n-propylsilyl derivatives.


Tremorine-oxotremorine-induced tremor, hypothermia and analgesia, and physostigmine toxicity, in mice after pretreatment with beta-adrenoceptor antagonists. The beta-adrenoceptor blockers propranolol, PhQA33 and LB-46 exhibited appreciable activity against tremorine-(TMN) and oxotremorine-(OTMN) induced tremor, whereas pronethalol, (+)-H56/28, (-)-H56/28, Kö-592 and L(+)-INPEA possessed weak action. The two beta-blockers, namely D,L(+/-)-INPEA and D(-)-INPEA acted as weak tremorgens. None of the above compounds suppressed the induced peripheral cholinergic phenomena; or possessed any central anticholinergic activity, as they were unable to afford protection against physostigmine-induced death. Propranolol, PhQA33 and LB-46 antagonized TMN-induced hypothermia and analgesia, but were inactive against OTMN-induced changes. A correlation of the beta-blocking and anti-tremor activity of these agents is unlikely.


The isolated cremaster muscle preparation and (external) spermatic nerve-cremaster muscle preparation of the guinea-pig. The isolated cremaster muscle preparation and spermatic nerve-cremaster muscle preparation of the guinea-pig were studied in vitro to determine their suitability as pharmacological test models. The preparation was contracted by acetylcholine, carbachol, succinylcholine and decamethonium (pD2 values, 4-2, 5-3, 7-3 and 7-4, respectively) through an action on a curare-sensitive cholinoceptor. Lobeline and DMPP were ineffective. Nicotine contracted the muscle, but there was tachyphylaxis. Tubocurarine and hexamethonium presumably competitively antagonized acetylcholine (pA2 values, 7-3 and 5-8); lobeline was a non-competitive antagonist (pD'2 value, 6-4). Atropine and mecamylamine exerted a dualistic action against acetylcholine (final pD'2 values, 5-3 and 6-7, respectively). Tubocurarine, succinylcholine and decamethonium exhibited their typical action when tested with spermatic nerve-cremaster muscle preparation; the latter two drugs also produced muscle spasm. Hexamethonium was a weak blocker of neuromuscular transmission. Atropine, mecamylamine, lobeline and DMPP exhibited neuromuscular blocking activity; however, directly evoked muscle twitches were also notably affected. The cremaster muscle preparations seem to add usefully to the list of currently used in vitro tests, with the added advantage that a mammalian skeletal muscle model is used for simultaneous quantitative studies.


Mediation of prostaglandin E2 in the biphasic response to ATP of the isolated tracheal muscle of guinea-pigs. ATP, at a dose higher than 0-1 mug m1(-1), showed a biphasic action consisting of an initial increase followed by a gradual decrease of muscle tension in the isolated tracheal strip-chains of guinea-pigs. The pattern of this biphasic response to ATP varied with the level of basal tone of the preparation at the moment of application of ATP. A smiliar biphasic action was obtained by prostaglandin (PG) E2 among the various active substances studied including acetylcholine, histamine, catecholamines and various types of PG. Indomethacin (0-1 mug m1(-1) and aspirin (30 mug m1(-1)) completely abolished the ATP-induced inhibitory response observed in the presence of histamine (10 muM). Polyphloretin phosphate (100 mug m1(-1)) also significantly depressed the inhibitory response to ATP or PGE2. It is concluded that the response to ATP of the preparation is mediated by PGE2 released via the stimulation of its biosynthesis.


The effects of angiotensin I and angiotensin II on the isolated tracheal muscle of the cat. The effects of Asp1-beta-amide-Val5-angiotensin II (A II) and Asp1-Ile5-angiotensin I (A I) have been studied on the isolated continuously superfused cat tracheal muscle contracted by 5-hydroxytryptamine (5-HT). Both peptides have been shown to induce dose-dependent relaxation on this muscle. Similar effects have been obtained with synthetic bradykinin, prostaglandin E2 (PGE2), noradrenaline and histamine. The effects of bradykinin, A I and A II have been shown to be inhibited by aspirin but not by propranolol, metiamide, SC 19220 or a specific, competitive antagonist of A II. The relaxing effect of A I is not due to the conversion of decapeptide to octapeptide A II. The possible mechanism of the relaxing effects of A I and A II on the cat isolated tracheal muscle is discussed.


Anticholinergic activity of antipsychotic drugs in relation to their extrapyramidal effects. Antipsychotic drugs were evaluated with two indices of anticholinergic activity, mydriasis in mice in vivo and antagonism of carbamylcholine-induced contractions of guinea-pig tracheal strips in vitro. The drugs from most to least potent as oral mydriatic agents were mepazine, clozapine, thioridazine, promazine and chlorpromazine. Trifluoperazine, pimozide and haloperidol were inactive. These results were consistent with the hypothesis that anticholinergic activity of antipsychotic drugs is inversely related to their propensity to produce extrapyramidal effects in man. In vitro results appeared to predict the incidence of extraphyramidal effects less accurately than in vivo results.


Some factors influencing dissolution from salicylic acid-urea solid dispersions. Solid dispersion systems of salicylic acid-urea have been prepared using a fusion method. Two different methods of cooling the melt were employed, rapid cooling in liquid nitrogen and slow cooling in air. Differential scanning calorimetry and an X-ray diffraction technique were employed to investigate the nature of the fused mixture. Evidence was found of compound formation between the constituents. Dissolution rates of drug from non-disintegrating discs of solid dispersion systems were measured. Rapid cooling of the melt resulted in a much faster drug dissolution rate than from a corresponding mixture prepared by a slow cooling method. This phenomenon is explained by a difference in the sizes of drug particles produced under the different cooling conditions. Rapid cooling favoured the generation of many nucleation sites for the solid drug particles as the liquid was cooled, and hence many small particles were obtained. Conversely, slow cooling favoured the growth of the first few nuclei or solid drug particles, rather than the production of new nuclei, and hence large drug particles were obtained.


Aggregation of antiacetylcholine drugs in aqueous solution: micellar properties of some diphenylmethane derivatives. Light scattering methods have been used to examine the aggregation in aqueous solution of a series of antiacetylcholine drugs based on the diphenylmethane nucleus. The drugs investigated included adiphenine hydrochloride, piperidolate hydrochloride, benztropine mesylate, orphenadrine hydrochloride, chlorphenoxamine hydrochloride, lachesine hydrochloride, poldine methylsulphate, pipenzolate bromide, clidinium bromide, benzilonium bromide and ambutonium bromide. A micellar pattern of association was established for all compounds and critical micellar concentrations and aggregation numbers have been determined.


The crystallization behaviour of sulphathiazole. Previous investigations into the dissolution kinetics of sulphathiazole Form 1 have shown the process to be first order with respect to driving force. However, below 37 degrees first order kinetics did not apply, due to the surface reaction becoming the rate controlling factor. The present study examines the kinetics of crystallization over the temperature range 25 to 50 degrees. Crystallization is a process greatly dependent upon the high-entropy surface integration step which, unlike the dissolution process, is much more sensitive to crystal growth inhibitors. In the absence of inhibitors crystallization above 34 degrees follows first order kinetics but below 34 degrees the process changes to third order. Similar temperature effects on the dissolution of sulphathiazole have been previously reported.


Anomalies in some properties of powder mixtures. Mixtures of lactose and paracetamol and of lactose and oxytetracycline exhibit anomalous properties. The mean particle sizes, tensile strengths and flow properties of the mixtures are not proportionally intermediate between those of the constituents. The results are ascribed to changes that occur in the packing arrangements of the particles. These changes could have practical consequences in monitoring the progress of a mixing operation by measuring apparent particle size and in controlling the properties of granules, capsules and tablets prepared from the mixtures.


Fluorimetric assay of tetracycline mixtures. A simple and precise fluorimetric method is described for the simultaneous assay in plasma of a mixture containing chlortetracycline, demethylchlortetracycline and tetracycline. Assay within the therapeutic ranges of 0-5 mg litre(-1) is achieved by formation of strongly fluorescent aluminum/tetracycline complexes, without prior extraction or separation of the individual antibiotics. This is performed by determinations at the peak excitation and fluorescence emission wavelengths of each tetracycline chelate. The method can be similarly applied to the assay of oxytetracycline, rolitetracycline and minocycline.


Preliminary investigations of the metabolism and pharmacological activity of beta-hydroxytryptamines in mammals. beta-Hydroxytryptamine and beta-hydroxy-5-hydroxytryptamine were incubated with rat liver slices and oxidative deamination was established as the main route of metabolism: in both instances the corresponding indole-3-glycollic acids and indole-3-ethane diols were the major metabolites. However, the rates of deamination of beta-hydroxylated tryptamines, as measured manometrically, were found to be much slower than those of tryptamines nonhydroxylated in the side chain. The pharmacological activities of beta-hydroxylated tryptamines were tested in guinea-pigs on resistance of respiratory pathways, spontaneous respiration, electrocardiogram, blood pressure and isolated ileum, using tryptamine and 5-HT as reference substances. The effects of tryptamines hydroxylated in the side chain were in general similar to those of corresponding tryptamines but of much lower intensities; only in increasing the blood pressure was beta-hydroxytryptamine as active as tryptamine. The different reactions of these two groups of substances in the presence of some antagonists indicate that the receptors are probably not the same.


Differential effect of neuroleptic drugs on dopamine turnover in the extrapyramidal and limbic system. In gallamine-immobilized cats, the caudate nucleus and the nucleus accumbens septi were perfused by means of a push-pull cannula and dopamine was measured in the perfusate. Chlorpromazine (10 mg kg(-1)) and clozapine (20 mg kg(-1)), administered intravenously, enhanced the release of dopamine. The effect of chlorpromazine was similar in both regions whereas that of clozapine was more pronounced in the nucleus accumbens than in the caudate nucleus. Furthermore, in the rat, sulpiride, clozapine and thioridazine increased the homovanillic acid concentration in striatum and limbic system to a similar extent. However, following probenecid administration, the net effect of these drugs on homovanillic acid accumulation was more marked in the limbic system than in the striatum whereas haolperidol and chlorpromazine had a similar effect in the two regions. It is concluded that, in contrast to haloperidol and chlorpromazine, sulpiride, clozapine and thioridazine may preferentially affect the limbic dopaminergic transmission. This possibly accounts for the fact that sulpiride, clozapine and thioridazine display an antipsychotic action and yet cause less extrapyramidal side effects than haloperidol and chlorpromazine.


An in vitro method for the study of beta-receptor mediated effects on slow contracting skeletal muscle. The soleus muscle of a guinea-pig was dissected out under pentobarbitone anaesthesia and mounted on a holder in an organ bath containing Krebs solution. The tendon was attached to a force transducer and subtetanic contractions were evoked by electrical field stimulation (0.5 ms pulses at 10-12 Hz for 1.5 or 3 s every 22 s). The experiments were performed at 37 degrees. Terbutaline, a selective agonist at beta2-adrenoceptors, reduced the force of subtetanic contractions in a dose-dependent manner, the EC50 being 0.2 muM. The reduction was due to a lessened degree of fusion. The results conform to previous in vivo studies.


Effects of alpha-methyldopa on blood pressure in the anaesthetized dog. Intravenous infusion over 1 h of 20 mg kg(-1) of alpha-methyldopa produced hypotension in the anaesthetized dog. The magnitude of this effect was, however, inversely correlated with bodyweight. Either rapid intravenous injection of alpha-methyldopa or slow infusion of alpha-methyldopate, was less effective in lowering blood pressure than infusion of free alpha methyldopa in dogs of equivalent bodyweight. Infusion of alpha-methyldopa into a vertebral or internal carotid artery produced hypotensive responses but these were no greater and generally less than those obtained to intravenous infusion. alpha-Methyldopa was therefore capable of producing hypotension in the dog but no evidence was obtained for this being the result of an action within the brain.


The effect of cholinesterase inhibitors on the antimuscarinic effect of hemicholinium-3 (HC-3) in the rat. The effect of hemicholinium-3 (HC-3) on responses of the rat isolated bladder and ileum to acetylcholine and carbachol was investigated in the absence and presence of a number of anticholinesterases. Responses of the bladder to acetylcholine were potentiated by DFP, edrophonium, BW284C51 and physostigmine but were unaffected by the specific butyrylcholinesterase inhibitor iso-OMPA. Responses to carbachol were not potentiated by the anticholinesterases. HC-3 (1.7 X 10(-4) M) inhibited responses to carbachol without affecting those to acetylcholine. In the presence of physostigmine or DFP responses to acetylcholine were inhibited by HC-3 but no such inhibition was observed in the presence of BW284C51, edrophonium or iso-OMPA or a combination of the latter two anticholinesterases. Responses to carbachol were also inhibited to a greater extent in the presence of DFP. In the ileum, responses to acetylcholine were increased in the presence of DFP, edrophonium and physostigmine but were unaffected by iso-Ompa. responses to carbachol were not increased by any of the anticholinesterases. HC-3 (2.8 X 10(-4) M) inhibited responses to both acetylcholine and carbachol in the ileum and the degree of inhibition was not significantly altered by the presence of any of the anticholinesterases used. Although a weak anticholinesterase, HC-3 was also found to decrease the inhibitory action of physostigmine on the hydrolysis of acetylcholine by homogenates of rat ileum. A similar effect was noted with DFP but not with edrophonium. The results obtained do not support a prejunctional action for HC-3 in antagonizing responses to carbachol. It is concluded that in addition to an inhibitory action on the post-junctional muscarinic receptor HC-3 may interfere with the anticholinesterase activity of some cholinesterase inhibitors such as physostigmine and DFP but not edrophonium.


Rapid and sensitive colorimetric determination of cobalt(II). A highly selective and sensitive spectrophotometric determination of cobalt (II) was developed. 7-Nitroso-8-hydroxyquinoline-5-sulfonic acid sodium salt was used as the chromogenic reagent for color development. Although other metals form colored chelates with the ligand, it was possible to develop a selective method using McIlvaine's pH 8 citric acid-phosphate buffer. Under these conditions, iron(II), iron (III), copper (II), zinc (II), and manganese (II), minerals likely to be compounded with cobalt (II) in geriatric formulations, do not interfere with the precision of the method or the color development. Calcium (II) and magnesium (II) do not form colored chelates with the used ligand. Hormones, vitamins, and additives likely to be present along with the cobalt ion in pharmaceutical formulations do not interfere. The sensitivity is 0.37 mug of cobalt (II)/ml of sample solution.


High-pressure liquid chromatographic determination of the 15-epimer of dinoprost in bulk drug. The p-nitrophenacyl esters of dinoprost and its 15-epimer are well resolved using high-pressure liquid chromatography. Quantitation was achieved using the internal standard technique. The specially synthesized diphenylurea ester of cholic acid was found to be a model internal standard. Graphs of peak height ratios of the prostaglandin to the internal standard were linear with respect to the amount of prostaglandin injected, with the lower detection limit of the 15-epimer being about 0.5%. Data are presented that demonstrate the usefulness of this analytical technique in determining the concentration of the 15-epimer present during studies on the kinetics of decomposition of dinoprost.


Effect of edetate disodium and reduced glutathione on absorption of acetazolamide from GI tract of rats. The absorption of acetazolamide suspensions from in situ rat gastric and intestinal loop segments was studied. In 1 hr, 66.2 and 64.3% remained unabsorbed from the rat stomach and intestine, respectively. Although 1% (w/v) reduced glutathione and 1% (w/v) (24 mM) edetate disodium had no effect on gastric absorption, drug absorption from the rat intestine (1 hr) was increased 1.5 and 2 times, respectively. It was hypothesized that the relatively poor intestinal absorption was due primarily to the formation of a pH-dependent (pH 4.5-10), nonabsorbable complex between acetazolamide and carbonic anhydrase present in the gut and that reduced glutathione acted as an inhibitor to promote intestinal absorption. Equilibrium dialysis studies showed that reduced glutathion could reduce the fraction of drug bound to human carbonic anhydrase B by one-half when present in a molar ratio 10 times that of acetazolamide; edetate disodium had no effect on the in vitro binding. It was, therefore, assumed that edetate disodium promoted an increase in intestinal absorption by altering the permeability of intestinal epithelium. Based upon present experimentation, however, the alteration of intestinal epithelium by reduced glutathione cannot be ruled out.


Chlorpromazine metabolism VIII: blood levels of chlorpromazine and its sulfoxide in schizophrenic patients. A procedure was standardized for extracting chloropromazine and its sulfoxide from the blood and for applying a recently developed fluorometric assay method to determine blood levels of these two compounds in schizophrenic patients receiving chlorpromazine therapy. The described methodology opens avenues for performing bioavailability and generic equivalence studies in humans.


Effects of various hydrodynamic conditions on dissolution rate determinations. An automated potentiometric procedure was used in dissolution rate studies to determine the effects of various hydrodynamic conditions on dissolution rate determinations. Changes in the hydrodynamics of the system resulted from using various sizes and shapes of dissolution vessels. Dissolution rate constants for benzoic acid prills in distilled water at pH-stat 6.2 were used as a measure of the agitation intensities present in the different shaped vessels. Great variations in the dissolution rates occurred in vessels with the same diameter and stirrer blade position when the shapes of the bottom of the vessel were varied. A similar order of dissolution rates was obtained at 100 and 150 rpm for the individual vessels at various propeller heights. The order differed from one vessel to another, depending on the shape of the bottom (concave, convex, or flat) of the vessel. In some cases, a change in the type of bottom resulted in the opposite order of rates for vessels with the same diameter.


Determination of tinidazole in tablets by dc polarography. A polarographic method was developed for determining tinidazole in tablets. The substance is extracted from the sample with water, pH 6.8 Britton-Robinson buffer is added to an aliquot, and the solution is polarographed at the dropping mercury electrode versus the saturated calomel electrode. The polarographic wave is well developed and enables a fairly precise quantitative determination. The method of standard addition is used.


Spectrophotometric determination of diphenhydramine hydrochloride using dipicrylamine. A spectrophotometric procedure for the determination of diphenhydramine hydrochloride based on the reaction with dipicrylamine was developed. A yellow complex forms and is easily extractable by chloroform at pH 5. The mole ratio of diphenydramine hydrochloride to dipicrylamine in the complex is 1:3. The absorbance of the complex obeys Beer's law over the concentration range of 3-10 mug of diphenhydramine hydrochloride per ml of chloroform. This procedure can be carried out in the presence of other compounds without interference.


Application of trihydroxyindole reaction to methanesulfonanilides: fluorometric analysis for soterenol and mesuprine. Methanesulfonanilides, like soterenol and mesuprine, which are bioisosteric with adrenergic catecholamines, form fluorescent species when subjected to the trihydroxyindole reaction. Presumably, the fluorescence is due to adrenolutin-like species formed from aminochrome intermediates. Fluorescence was not induced in a relative of soterenol where a methyl group was added to the sulfonamido nitrogen, a fact that suggests the presence of a quinoid intermediate in the reaction scheme of soterenol. A ring isomer of soterenol, where the methanesulfonamido and hydroxyl groups were interchanged, produced only about 5% as much fluorescence response as soterenol. The sterenol counterparts to isoproterenol and isoproterenol sulfonic acid did not produce fluorescence when treated like soterenol. This finding and the fact that response was linear with concentration for soterenol and mesuprine suggest that a fluorometric analysis could be developed for these methanesulfonanilides.


Continuous dissolution rate determination as a function of the pH of the medium. A method was developed for varying the pH of the medium during dissolution rate studies of timed-release tablets with the aid of compressed, totally soluble, alkaline powder mixtures. Commercial as well as experimental timed-release capsules or tablets were used as models, and dissolution rates were determined at pH 1.1, 2.4, and 7.4. The system can be applied to other pH values or other variations of the dissolution medium (e.g., ionic strength) to: (a) correlate in vitro release rates with bioavailability data, (b) discriminate between alternative formulations during dosage form development, or (c) serve as a selective control procedure for a series of sustanined-release dosage forms.


Controlled flocculation of coarse suspensions by colloidally dispersed solids I: Interaction of bismuth subnitrate with bentonite. Deflocculated suspensions of coarse powders tend to cake as the individual particles settle out and form compact, cohesive sediments. Limited flocculation results in looser sediments because the settled-out flocs incorporate large amounts of the liquid suspending medium. Controlled flocculation of bismuth subnitrate suspensions was achieved by the addition of small amounts of bentonite. The interaction of the coarse, positively charge bismuth subnitrate particles in aqueous suspension with negatively charged, colloidally dispersed bentonite was investigated by measuring electrophoretic mobility, sedimentation volume, and viscosity. Gradual addition of bentonite dispersion to bismuth subnitrate suspensions first reduced the zeta-potential of the bismuth subnitrate particles from +28 mv to zero, then inverted it, and finally caused it to level off at -20 mv for bismuth subnitrate-bentonite weight ratios below 200. Owing to the much greater specific surface area of bentonite, the surface of the bismuth subnitrate lath-shaped crystals was completely covered by 0.5% of its weight in clay platelets. Adhesion was promoted by electrovalences between surface bismuthyl ions and cation-exchange sites of the clay and by secondary valences. The charge neutralization of bismuth subnitrate by bentonite was a heterocoagulation process: the addition of small amounts of the clay flocculated the bismuth subnitrate suspensions and eliminated caking. While the zeta-potential of the bismuth subnitrate particles leveled off when their surface was saturated with bentonite platelets, sedimentation volume and viscosity continued to increase when the clay concentration was increased further while maintaining the bismuth subnitrate concentration constant. The excess, nonadsorbed bentonite formed the characteristic house-of-cards structure, incorporating the bentonite-coated bismuth subnitrate particles as cornerstones.


Urinary excretion of probenecid and its metabolites in humans as a function of dose. A GLC assay was used to study the excretion of probenecid and its metabolites in the urine of human subjects following oral doses of 0.5, 1, and 2 g. From 75 to 88% of the  dose was found in the urine. The major metabolite, probenecid acyl glucuronide, accounted for 34-47% of the dose. Approximately equal amounts (10-15%) of the mono-N-propyl, secondary alcohol, and carboxylic acid metabolites were excreted in the unconjugated from with only traces in the conjugated form. The primary alcohol metabolite was not found in measurable amounts. The terminal half-lives for excretion of all metabolites were in the range of 4-6 hr, were independent of dose, and were limited by their rates of formation. A prolonged time course of excretion of the metabolites, particularly at higher doses, suggests that probenecid, being poorly soluble in water, precipitates from solution in the GI tract, forming a depot of drug from which absorption is dissolution rate limited. The urinary excretion of unchanged probenecid, which accounts for 4-13% of the dose, is dependent on both the pH and flow rate of urine.


Nitrofurantoin solubility in aqueous urea and creatinine solutions. Experiments were carried out to determine the effect of urea and creatinine on the solubility of nitrofurantoin in water at different temperature and pH conditions. The addition of urea to aqueous media increased nitrofurantoin solubility up to a maximum concentration level and then decreased solubility at higher urea concentrations. The amount of urea needed to bring about maximum nitrofurantoin solubility was dependent on temperature and ranged between 1.75 and 2.50%. Spectral studies suggest a possible interaction between urea and nitrofurantoin molecules. Nitrofurantoin solubility increased with an increasing creatinine concentration ranging from 0.05 to 1.6%. Spectral studies indicate a strong interaction between creatinine and nitrofurantoin molecules in solution. The combined effect of urea and creatinine of the solubility of nitrofurantoin could account for the absence of crystalluria with this drug, even though unusually high concentrations in urine have been reported.


Influence of pH and route of injection on acute toxicity of tetracycline in mice. Some LD50 determinations for tetracycline hydrochloride in mice were carried out over a range of pH values, using both the intraperitoneal and the subcutaneous routes of injection. Depending on the pH of the formulation, either water or a solvent system of water and 60% (v/v) propylene glycol was employed thus ensuring complete solution of the drug at all pH values tested. Higher LD50 values were obtained with the subcutaneous route than with the intraperitoneal route. With either route of administration, there was a trend toward lowest LD50 value occurrence at the isoelectric pH of tetracycline. Actual statistical significance was achieved for the intraperitoneal route only when LD50 values obtained at the isoelectric pH were compared with either of the more acidic pH values.


Monoamine oxidase inhibition and the induction of ponto-geniculo-occipital wave activity by reserpine in the cat. Reserpine induces ponto-geniculo-occipital wave activity similar to that seen in the cat during the rapid eye movement phase of sleep. This action of reserpine was blocked by the monoamine oxidase inhibitors, pheniprazine, harmaline and clorgyline, but not deprenyl or its demethylated analog. After a single dose (20 mg/kg) of either pheniprazine or harmaline, the time course of antagonism of the effect of reserpine was in good correspondence with inhibition of monoamine oxidase but not with restoration of the serotonin content of several regions of the brain. Harmaline had a reversible effect while that of pheniprazine persisted for weeks. Clorgyline in low doses (0.5-1.0 mg/kg), at which it is a specific inhibitor of type A monoamine oxidase, antagonized the action of reserpine for over 2 weeks. Deprenyl and its demethylated analog failed to suppress the reserpine-induced waves even at 10 times the dose of clorgyline when they should have completely inhibited type B monoamine oxidase. In contrast to antagonism of reserpine, suppression of rapid eye movement sleep by these monoamine oxidase inhibitors could be temporally dissociated from their inhibition of the enzyme. It is concluded that suppression of the induction of ponto-geniculo-occipital waves by reserpine after administration of the monoamine oxidase inhibitors is a specific effect of these drugs and is related to inhibition of type A monoamine oxidase. Suppression of rapid eye movement sleep is probably a nonspecific effect and not related to inhibition of the enzyme.


Further studies on angiotensin tachyphylaxis. Rabbit aorta and rat fundus do not display tachyphylaxis to angiotensin II (Asp1)angiotensin II) at 37 degrees C. These tissues do, however, display tachyphylaxis to [Sar1]angiotensin II (N-methylglycine 1 angiotensin II) and [dimethylglycine angiotensin II, respectively, at this temperature. At 22 degrees C, both tissues displayed rapid tachyphylaxis to [Asp1]angiotensin II which was reversed on prolonged incubation in peptide-free medium. At 37 degrees C, but at a lower than normal pH (pH 6.5 instead of 7.4), the rabbit aorta developed rapid tachyphylaxis to [Asp1)angiotensin II; this effect also was reversed on prolonged incubation in peptide-free medium. The rat uterus, at 30 degrees C, displayed tachyphylaxis to [Asp1]angiotensin II and analogs in order of rate of onset: [Sar1]angiotensin II greater than [Asp1]angiotensin II greater than [des-Asp1]angiotensin II. Over half of the individual uterine strips tested displayed no tachyphylaxis to [des-Asp1]angiotensin II. However, at 22 degrees C tachyphylaxis to all three compounds was rapid on onset and was reversed on prolonged incubation. The results indicate a correlation between affinity of the peptide analog for angiotensin receptor and its ability to induce tachyphylaxis. A possible involvement of membrane-bound enzymes in the onset and duration of tachyphylaxis is suggested.


Studies on adrenal phenylethanolamine N- methyltransferase (PNMT) with S K & F 64139, a selective inhibitor. SK&F 64139 is a potent, reversible inhibitor of phenylethanolamine N-methyltransferase; its IC50 concentration in our standard assay system was 1 X 10(-7) M. Kinetically, the compound is a competitive inhibitor with respect to norepinephrine but is uncompetitive when S-adenosylmethionine is the variable substrate. In contrast to a previously reported compound (SK&F 7698), the drug is only a weak alpha receptor antagonist (KB = 6 X 10(-6) M). In both the rat and squirrel monkey, SK&F 64139 produced dose-dependent decreases in the adrenal epinephrine content coupled with stoichiometrically equivalent increases in the norepinephrine pool(s). Further evidence for in vivo phenylethanolamine N-methyltransferase inhibitory activity was that the drug markedly inhibited the conversion of a tracer dose of 3H-norepinephrine to 3H-epinephrine in the rat adrenal gland after unit oral doses as low as 5 mg/kg.


The effects of metoclopramide in modifying the response of isolated guinea-pig ileum to various agonists. Metoclopramide has a dual effect on intestinal smooth muscle. Low concentrations of metoclopramide cause potentiation of the responses to substance P, acetylcholine, histamine and barium chloride on the guinea-pig ileum. Higher concentrations produce a depression of smooth muscle responses which is characteristic of the tertiary amine local anesthetics. Neural pathways are involved in the mechanism of potentiation, since the enhancement of the responses to the agonists is abolished by tetrodotoxin. Atropine partially antagonizes the potentiating effect of metoclopramide implying that activation of muscarinic receptors is a contributing factor, but this does not fully explain the potentiation.


Statistical parameters of transmitter release at frog neuromuscular junctions treated with guanidine or tetraethylammonium. Transmitter release at Mg++ -depressed frog neuromuscular junctions can be described using binomial statistics. The number of quanta (m) released by the nerve terminal action potential is directly proportional to the mean probability (p) that a quantum will be released and the number of quanta available for release (n). Guanidine or tetraethylammonium (TEA) increased m and n, but had no effect on p. At junctions depressed by d-tubocurarine, both compounds enhanced the amplitude of the initial end-plate potential, caused an accelerated rate of fade of end-plate potential amplitudes and raised the steady-state level of end-plate potential amplitude. This finding was interpreted to mean that guanidine and TEA increased transmitter mobilization and may be related to the increase by the compounds of the parameters n. If so, then the data support the idea that n represents the number of quanta available for release rather than the number of transmitter release sites in the terminal membrane. Neither compound affected the power relationship between [Ca++]o and transmitter release. When plotted on a double logarithmic basis, the slope of the line relating [Ca++]o to increased transmitter release was 3.7, a relationship not altered by the drugs. Thus, the compounds had no effect on the fundamental interaction between Ca++ and transmitter release sites. In contrast to [Ca++]o, the power relationship between increased transmitter release and the concentration of drug in the bathing solution was 0.69 for guanidine and 0.84 for TEA. Because of this finding, it was concluded that the compounds increased transmitter release by mechanisms other than or in addition to increasing Ca++ conductance.


Effect of fatty acids on the disposition of ammonia. Subcoma doses of fatty acids and ammonium salts injected intraperitoneally at the same time into rats or cats act synergistically to produce coma. Under these circumstances, the blood ammonia, is more than double that when the NH4+ is given alone. After these observations a rat liver homogenate system was utilized to study the effect of fatty acids on ammonia utilization in urea, glutamate and glutamine synthesis in vitro. Acetylglutamate-catalyzed urea synthesis was completely inhibited by 45 mM octanoate and was depressed 46% by 9.5 mM octanoate. Citrulline synthesis was similarly inhibited 86 and 28%, respectively. The concentration of liver octanoate at the moment of occurrence of coma after an in vivo injection was approximately 10 mM. The inhibitory effect of fatty acids on the utilization of NH4+ in the urea cycle was greater the longer the fatty acid chain. The critical step in this interference with ammonia metabolism was the inhibition of carbamyl phosphate synthetase. Argininosuccinate synthetase activity was also inhibited to a lesser degree, but ornithine transcarbamylase, argininosuccinate lyase and arginase were unaffected. Glutamate dehydrogenase was likewise inhibited in liver (83%) and brain (43%) by 13 mM octanoate, whereas glutamine synthetase was unaffected. Thus, the two main processes whereby ammonia is metabolized were inhibited by fatty acids at concentrations that exist pathologically, which accounts, at least in part, for the rise in blood ammonia in vivo.


Microsomal reductive glycosidase.

Microsomal reductive glycosidase. Rat liver microsomes contain a phenobarbital inducible, NADPH dependent, reductive glycosidase capable of cleaving several anthracycline antibiotics, including adriamycin and daunorubicin, to deoxyaglycone products. The pH optimum for the reaction ranges from 7 to 7.4, and no metal requirements are noted. Molecular oxygen reversibly inhibits the microsomal enzyme greater than 95% at 20% oxygen partial pressure. Carbon monoxide, SKF 525A and sulfhydryl reagents are not inhibitory to the reaction, but the enzyme is sensitive to Cu++ and Zn++. Since the intact glycoside is necessary for conversion to the deoxyaglycone and a possible intermediate hydroxylated aglycone is not reduced to the deoxyaglycone, a concerted reaction mechanism is proposed. The reductive glycosidase activity is also present in rat brain, kidney and other tissues. Sensitivity of this enzyme to molecular oxygen suggests a possible regulatory role for the enzyme in vivo.


Neutral protease activity and erythropoietin production in the rat after cobalt administration. The generation of erythropoietin purportedly involves the interaction of an enzyme, of renal origin, and a circulating plasma protein substrate. Cobalt, which has the capacity to stimulate erythropoietin formation, was evaluated for its effects on renal and plasma protease activity; in addition, the nature of the erythrogenic mechanism was investigated. Rats receiving one s.c. injection of cobalt demonstrated significant increases in activity of renal cathepsins A and B. The activity of a plasma protease was also elevated in rats after cobalt treatment. Cathepsins A and B and plasma protease were sensitive to the same enzymatic inhibitory agents, and all three enzymes demonstrated activity patterns similar to that of the renal erythropoietic factor [enzyme(s) presumed responsible for generating erythropoietin] in rats treated with cobalt. These data may help to elucidate the mechanism of erythropoiesis.


Cholestyramine-induced inhibition of salicylazosulfapyridine (sulfasalazine) metabolism by rat intestinal microflora. The effect of multiple oral administration of the hypocholesterolemic agent cholestyramine (a strongly basic anion-exchange resin) on the metabolism of salicylazosulfapyridine by microflora present in the colon and cecum was assessed in conventional rats by following the time course of salicylazosulfapyridine and its metabolites in the urine and feces. The intestinal metabolism of salicylazosulfapyridine (a single 100 mg/kg oral dose), which involves reduction of the azo linkage by bacterial azo reductases and the liberation of sulfapyridine and 5-aminosalicylic acid (potential active metabolites of the drug), was markedly inhibited by the resin (250 mg/kg oral doses at -2, +2 and +6 hours), resulting in an enhanced fecal excretion of intact salicylazosulfapyridine. The existence of a rank-order correlation between the in vitro binding of salicylazosulfapyridine, sulfapyridine and 5-aminosalicylic acid to the resin and their fecal excretion pattern in resin-treated animals suggests that a direct cholestyramine-salicylazosulfapyridine interaction occurred within the intestinal tract and that in the bound state, the azo bond of the drug was inaccessible to bacterial azo reductases. These findings suggest that chronic oral administration of cholestyramine to patients with ulcerative colitis who are receiving salicylazosulfapyridine could result in a significant reduction in the absorption and metabolism of the drug and hence, in its therapeutic efficacy.