Extract of kava (Piper methysticum) and its methysticin constituents protect brain tissue against ischemic damage in rodents. The purpose of the present study was to test whether kava extract and its constituents kawain, dihydrokawain, methysticin, dihydromethysticin and yangonin provide protection against ischemic brain damage. To this end, we used a model of focal cerebral ischemia in mice and rats. Ischemia was induced by microbipolar coagulation of the left middle cerebral artery (MCA). To quantify the size of the lesion in mice, the area of the infarct on the brain surface was assessed planimetrically 48 h after MCA occlusion by transcardial perfusion of carbon black. In the rat model infarct volume was determined 48 h after MCA occlusion by planimetric analysis and subsequent integration of the infarct areas on serial coronal slices. Compounds were administered i.p., except the kava extract, which was administered orally. The effects of the kava extract and its constituents were compared with those produced by the typical anticonvulsant, memantine. The kava extract, methysticin and dihydromethysticin produced effects similar to those of the reference substance memantine. The kava extract (150 mg/kg, 1 h before ischemia) diminished the infarct area (P less than 0.05) in mouse brains and the infarct volume (P less than 0.05) in rat brains. Methysticin, dihydromethysticin (both 10 and 30 mg/kg, 15 min before ischemia) and memantine (20 mg/kg, 30 min before ischemia) significantly reduced the infarct area in mouse brains. All other compounds failed to produce a beneficial effect on the infarct area in mouse brains. In conclusion, the kava extract exhibited neuroprotective activity, which was probably mediated by its constituents methysticin and dihydromethysticin.


The effect of intravenous recombinant human renin on blood pressure in pithed spontaneously hypertensive rats. The effect of highly purified recombinant human renin (rh-renin), expressed in Chinese hamster ovary cells, on mean blood pressure (MBP) was evaluated in pithed spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). Intravenous bolus injection of rh-renin produced dose-dependent increases in MBP in pithed SHR and WKY. The pressor response to rh-renin in pithed SHR was about 3 times as potent as that in pithed WKY. Intravenous infusion of rh-renin produced dose-dependent progressive increases in MBP during the first 40 min, reaching plateaus and thereafter MBP was maintained up to 120 min. This hypertensive response to rh-renin was antagonized by renin inhibitors, YM-21095 and KRI-1314, which inhibited the reaction between rh-renin and tetradecapeptide competitively, with Ki values of 5.1 x 10(-10) and 4.3 x 10(-9) M, respectively. In rh-renin-infused pithed SHR, the hypotensive effect of YM-21095 was 37 times as potent as that of KRI-1314. These results suggest that rh-renin can stimulate the rat renin-angiotensin system, thereby producing hypertension. Moreover, the rh-renin-infused rat model could be useful to evaluate the effect of renin inhibitor.


Inhibition of uptake of 1-methyl-4-phenylpyridinium ion and dopamine in striatal synaptosomes by tobacco smoke components. To determine whether the attenuation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neurotoxicity by tobacco smoke exposure is caused by inhibition of the neuronal uptake of 4-phenylpyridinium ion (MPP+), various tobacco components and a smoke extract were tested for inhibitory activity in striatal synaptosomes. A dimethylsulfoxide extract of tobacco smoke filtrate was found to inhibit the uptake of MPP+ and dopamine. These results suggest that inhibition of the neuronal dopamine uptake mechanism may account for the protective effects of smoke exposure on MPTP-induced neurotoxicity.


Effect of aldose reductase inhibitor on the inhibition of platelet aggregation induced by diabetic rat plasma. Plasma isolated from streptozotocin-induced diabetic rats inhibited platelet aggregation. Treatment of diabetic rats with an aldose reductase inhibitor, ONO-2235, which did not improve hyperglycemia, prevented the antiaggregating activity of plasma as did insulin treatment. Both treatments reduced the elevated sorbitol concentrations in erythrocytes. Although the factor(s) accounting for the antiaggregatory effect of diabetic plasma has not yet been characterized, it is possible that the accumulation of sorbitol in erythrocytes is related to the generation of the factor(s).


Chronic morphine treatment causes down-regulation of spinal adenosine A1 receptors in rats. Recent studies suggest that the release of adenosine in the spinal cord may be a significant component of the morphine antinociceptive action. We wanted to know whether the spinal adenosine system is involved in morphine tolerance. Animals were rendered tolerant to morphine, and A1 adenosine receptor binding activity was measured. Treating Sprague-Dawley rats with multiple, increasing doses of morphine i.p. for 6 days resulted in an about 10-fold increase in the median antinociceptive dose (AD50) of morphine to elicit an antinociceptive response. On the other hand, this treatment also caused a 4 to 5-fold increase in the AD50 of cyclopentyladenosine (CPA). When A1 adenosine receptor binding was determined by using [3H]cyclohexyladenosine ([3H]CHA) a significant decrease in binding (P less than 0.05) in the spinal cord but not in the cortex was observed. Scatchard analysis of the [3H]CHA saturation binding data revealed a decrease in Bmax values (from 185.5 fmol/mg to 110.2 fmol/mg) and no significant change in Kd values.


Antihypertensive and hormonal activity of MK 954 in spontaneously hypertensive rats. MK 954 (DuP 753), a recently developed angiotensin II (Ang II) receptor antagonist, was administered orally for 2 weeks to spontaneously hypertensive (SHR) and Wistar-Kyoto rats (WKY). Whereas the basal levels of plasma Ang II were lower in SHR than in WKY, treatment with MK 954 markedly reduced blood pressure in SHR but not in WKY. Plasma renin activity, Ang I and Ang II were increased, while plasma aldosterone was decreased in both strains. These results no only indicate therapeutic efficacy of this agent in the chronic treatment of human hypertension, but also support the idea that the renin-angiotensin system plays an important role in the control of blood pressure in SHR.


Release of immunoreactive [Met5]enkephalin and cholecystokinin from the spinal cord by beta-endorphin administered intraventricularly in the pentobarbital-anesthetized rat. Beta-endorphin (4 micrograms) injected i.v.t. induced the release of immunoreactive [Met5]enkephalin and sulfated cholecystokinin octapeptide (CCK-8s) from the spinal cord in the pentobarbital-anesthetized rat. The inhibition of the tail-flick response induced by beta-endorphin given i.v.t. was antagonized by i.t. injection of CCK-8s and potentiated by i.t. administration of the antibody to CCK-8s. The findings give support to the hypothesis that endogenous CCK-8s in the spinal cord may play a negative feedback role in modulating beta-endorphin-induced antinociception.


Striatal dopamine D1-like receptors have higher affinity for dopamine in ethanol-treated rats. Experiments were carried out with brain tissues of ethanol-experienced (long-term ethanol intake but withdrawn) vs. ethanol-naive animals. The in vitro 3H antagonist binding of [3H]SCH 23390 and of [3H]spiperone to striatal dopamine D1- and D2-like receptors revealed no significant changes in KD and Bmax values. Displacement of the 3H antagonist binding by dopamine indicated high- and low-affinity states, which also showed no significant alteration at the D2-like receptor but a 5-fold increase of dopamine affinity at the high-affinity state of the D1-like receptor of the ethanol-experienced rats.


Effects of omega-3, omega-6 and omega-9 fatty acids on vascular smooth muscle tone. The comparative effects of omega-3, omega-6 and omega-9 fatty acids on vascular smooth muscle tone were investigated. Docosahexaenoic acid (1-255 microM) and eicosapentaenoic acid (31-255 microM) inhibited phenylephrine-induced contractions, (8-63%) and (20-65%), respectively, which were not altered by indomethacin, NDGA, or by removal of the endothelium. Linoleic acid (18:2n6) and arachidonic acid (20:4n6) also induced significant relaxation. Therefore, fatty acid-induced relaxation of the rat aorta is specific to polyunsaturated fatty acids, 20:5n3, 22:6n3, 18:2n6 and 20:4n6.


Transmural pressure inhibits nitric oxide release from human endothelial cells. We examined the effect of transmural pressure on histamine-stimulated nitric oxide release from cultured endothelial cells prepared from human umbilical cord veins. PO2 and pH were kept constant throughout the experiments. Various levels of transmural pressure and atmospheric pressure (40, 80, 120 and 160 mm Hg) were applied. Nitric oxide release was inhibited in a pressure-dependent manner. The inhibitory effects were reversible, and nitric oxide had no effect on the morphology of the cells. Our results suggest that transmural pressure-mediated inhibition of nitric oxide release contributes to pressure-induced vasoconstriction and reduced endothelium-dependent relaxation in patients with hypertension.


Effect of naloxone-precipitated morphine withdrawal on noradrenaline release in rat hippocampus in vivo. Here we investigated the effect of naloxone-precipitated morphine withdrawal on the release of noradrenaline in hippocampus of the anaesthetised rat. Naloxone (1 mg/kg i.p.) injected 3 and 24 h, but not 3 weeks, after eight daily injections of morphine, induced an immediate increase in hippocampal noradrenaline release. This striking effect of naloxone was markedly attenuated by pretreatment with clonidine (0.1 mg/kg s.c.). These findings provide direct evidence for a marked release of noradrenaline in hippocampus during opiate withdrawal in vivo.


C-type natriuretic peptide-22 differentiates between natriuretic peptide receptors in rat choroid plexus and subfornical organ. Atrial natriuretic peptide (ANP) receptors in the rat choroid plexus and subfornical organ were characterized with C-type natriuretic peptide-22 (CNP-22) and 125I-Tyr0-CNP-22. The receptor autoradiographic method we used revealed that 125I-Tyr0-CNP-22 specifically bound to the choroid plexus, but only slightly to the subfornical organ, areas densely labeled with 125I-alpha-rat ANP (125I-rANP). CNP-22 significantly inhibited 125I-rANP binding to the choroid plexus; however, the peptide did not affect 125I-rANP binding to the subfornical organ. Thus, the characteristics of ANP receptors in the rat choroid plexus and subfornical organ are probably different.


Inhibition by azelastine of the effects of platelet-activating factor in lungs from actively sensitised guinea-pigs. The effect of azelastine on platelet-activating factor (PAF)-induced bronchoconstriction and mediator release in isolated lungs from actively sensitised guinea-pigs was investigated. Guinea-pigs were actively sensitised with two s.c. injections of 10 micrograms ovalbumin in 1 mg Al (OH)3 at a 2-week interval. One week after the second injection, the lungs were removed and challenged intra-arterially with PAF or arachidonic acid. In some experiments lungs from non-immunised guinea-pigs were injected with PAF or histamine. Bronchoconstriction, the release of thromboxane (TX)B2 or leukotriene (LT)-like material and the histamine content of the effluent were evaluated. Azelastine was given s.c. at 10 or 100 micrograms/kg, 4 h before lung removal. Azelastine (100 micrograms/kg) did not inhibit PAF-induced bronchoconstriction and mediator release from lungs from non-immunised guinea-pigs. In contrast, the hyperresponsiveness to 1 ng PAF observed in lungs from actively sensitised animals was dose dependently inhibited by azelastine. Azelastine did not reduce the histamine-induced bronchoconstriction and consequent TXB2 release from lungs from immunised guinea-pigs, indicating that the protective effect exerted against hyperresponsiveness to PAF was not due to histamine antagonism. Azelastine also reduced arachidonic acid-induced bronchoconstriction and LT-like material release from sensitised lungs, regardless of the presence of indomethacin. These results suggest that inhibition of lung hyperresponsiveness to PAF by azelastine may result from an interference with leukotriene synthesis.


Effects of systemic morphine on diffuse noxious inhibitory controls: role of the periaqueductal grey. The effects of systemic morphine (1 mg/kg i.v.) on diffuse noxious inhibitory controls (DNIC) were studied in both sham-operated animals and those with quinolinic acid-induced lesions of the periaqueductal grey (PAG). DNIC acting on convergent neurones in the dorsal horn of the spinal cord were similar in the sham-operated and lesioned animals. However, following morphine injection, DNIC were blocked in the sham-operated but not in the PAG-lesioned animals. It is concluded that, although the PAG is not directly involved in the supraspinal loop subserving DNIC, it can modulate these controls. In addition, as naloxone reversed the effects of morphine in the control group but reduced DNIC in the PAG-lesioned animals, it is suggested that more than one opioidergic system is involved in DNIC.


Effect of SR 27417 on oedema formation induced in rabbit skin by platelet-activating factor or antigen. SR 27417, the first member of a newly developed series of platelet activating factor (PAF) antagonists inhibited in a dose-dependent manner PAF-induced oedema formation in rabbit skin when administered i.d. premixed with PAF (ED50 = 3.3 +/- 0.15 pmol/site i.d. (intradermally) (n = 5). The effect of SR 27417 was over 660 times more potent than that of the triazolothienodiazepine, WEB-2086 (ED50 = 5.4 +/- 0.71 nmol/site i.d.) (n = 5). SR 27417 protected rabbits from PAF-induced plasma extravasation with an ED50 value of 16 +/- 3 micrograms/kg when given i.v. 1 h before PAF challenge. It was also effective when given p.o. 3 h before PAF i.d. administration (ED50 = 0.18 +/- 0.07 mg/kg p.o.) (n = 5). This effect of SR 27417 was selective for PAF since inflammatory responses induced by other mediators (bradykinin, histamine, N-formyl-L-methionyl-L-leucyl-L-phenyl-alanine, leukotriene B4) were not affected. After i.v. or oral administration (1 and 5 mg/kg respectively) SR 27417 had an extended duration of activity (between 72 and 96 h). In presensitized rabbits, SR 27417 premixed with the allergen inhibited dose-dependently allergen-induced plasma exudation (ED50 = 12 +/- 0.08 nmol/site i.d.) (n = 5) (vs. 850 +/- 98 nmol/site (n = 5) for WEB-2086). Similarly, i.v. injection of SR 27417 (1 mg/kg i.v.) inhibited allergen-induced vascular permeability. These results confirm that PAF plays a major role in the development of cutaneous anaphylaxis and that SR 27417 may be an effective prophylactic drug.


Protective effect of bepridil against veratrine-induced contracture in rat atria. In isolated stimulated rat atria, superfusion with veratrine caused a marked contracture (VIC) which was absent in calcium-free medium and which was inhibited by tetrodotoxin (IC50VIC of 1.38 microM). Lowering the extracellular calcium concentration from 2.5 to 0.5 or 0.1 mM reduced the veratrine-induced contracture and delayed its onset. Superfusion of bepridil (1-10 microM) for 60 min before and during veratrine exposure markedly slowed the onset of contracture, reduced the maximum response (IC50VIC = 2.11 microM) and facilitated recovery upon washout of the alkaloid. The direct negative inotropic effect (NIE) of bepridil (IC50NIE = 10.96 microM) resulted in an VIC/NIE ratio of 5.19 for this drug. The protective effects of bepridil were rate-independent and were not modified by the presence of atropine (1.4 microM) and propranolol (0.3 microM) in the medium. Diltiazem, verapamil and nifedipine only reduced veratrine-induced contracture at concentrations much higher than those producing a negative inotropic effect, giving them negative NIE/VIC ratios of 0.31, 0.08 and 0.08 respectively. Like bepridil, flunarizine had a positive NIE/VIC ratio (15.87, IC50VIC = 3.71 microM). The lack of effect of the quaternary derivative of bepridil CERM 11888 indicated that intracellular sites of action may be involved in the activity of bepridil on veratrine-induced contracture. Given that veratrine-induced changes may mimic some of the pathological changes occurring in ischaemia, the results suggest that bepridil and flunarizine may be more effective than L-type, slow calcium ion-channel blockers in protecting against calcium overload during ischaemia and reperfusion injury.


Blockade by trifluoperazine of a Ca(2+)-activated K+ channel in rat hippocampal pyramidal neurons. The effects of trifluoperazine, a phenothiazine derivative, on the large-conductance Ca(2+)-activated K+ channel (BKCa) in dissociated rat hippocampal pyramidal neurons were examined using the inside-out configuration of the patch-clamp technique. The BKCa was activated by 12.6 microM Ca2+ on the internal surface of the membrane patch. The single channel conductance of the BKCa was 244 +/- 17.5 pS (n = 10) in symmetrical solutions of 150 mM K+. Trifluoperazine, applied on the internal surface of the membrane, decreased the open probability of the channel without changing the single channel conductance. The reduction in the open probability was well described by a block of the open state of the channel in a simple sequential model. The apparent dissociation constant (KD) for the reduction was calculated to be 1.4 microM and the Hill coefficient 0.69 at +20 mV. The inhibition was voltage dependent, being more pronounced at depolarized voltages. The voltage dependence enabled us to estimate that the binding site for the agent in the channel lies about half way across the membrane electrical field. It is concluded that trifluoperazine blocks the open state of the BKCa, which is known to provide an outward current for repolarization and afterhyperpolarization of the neuronal action potential. This may result in a decrease in spike intervals during burst firing of neurons.


Antagonism by beta-eudesmol of neostigmine-induced neuromuscular failure in mouse diaphragms. beta-Eudesmol, a sesquiterpenol extracted from a Chinese herb, Atractylodes lancea, at 10-80 microM, did not affect muscle action potentials, miniature and evoked endplate potentials and acetylcholine-induced depolarization in the presence or absence of neostigmine in mouse phrenic nerve-diaphragms. However, the tetanic fade, muscle fasciculation and twitch potentiation induced by neostigmine were effectively antagonized by 20 microM beta-eudesmol. When trains of pulses were applied to the nerve in the presence of neostigmine, beta-eudesmol reduced the incidence of explosive depolarization of the endplate from 95% to 35-67% of junctions, and shortened the duration when it occurred. Moreover, both the maximal and steady-state depolarizations during repetitive stimulation were reduced while the amplitudes of steady-state endplate potentials were increased. The results suggest that beta-eudesmol antagonized neostigmine-induced neuromuscular failure mainly by a presynaptic action to depress the regenerative release of acetylcholine during repetitive stimulation. The mechanism of antagonism is obviously not tubocurarine-like and it is unrelated to desensitization of acetylcholine channels.


Characterization of antigastrin activity in vivo of CR 2194, a new R-4-benzamido-5-oxo-pentanoic acid derivative. The antigastrinic activity, in vivo, of CR 2194 (R-4-(3-chlorobenzamido)-5-(8-azaspiro[4.5]decan-8-yl) -5-oxo pentanoic acid) was assessed in various animal species. CR 2194 antagonized pentagastrin-stimulated gastric acid secretion in the rat (ID50 = 11 mg/kg i.v.), dog (ID50 = 5.9 mk/kg i.v. or 28.8 mg/kg os) and cat (ID50 = 15.5 mg/kg i.v.). CR 2194, in the cat, inhibited both competitively and non-competitively the gastric acid secretion stimulated with increased doses of pentagastrin, with a pA2 of 4.89. In the rat and in the dog the antagonism seemed to be non-competitive and the respective pD'2 calculated were 4.54 and 4.42. The interaction of CR 2194 with the gastrin receptors appeared reversible, as demonstrated by the return to normal values of the acid output after the conclusion of the i.v. infusion, during pentagastrin continuous stimulation in the dog. The antigastrin activity was specific: CR 2194 was unable to antagonize the gastric acid secretion stimulated by carbachol or histamine in the rat up to the dose of 100 mg/kg. CR 2194 was effective to antagonize the gastric acid secretion stimulated by gastrin release after meal ingestion in the Heidenhain pouch dog model. The ID50 calculated was 2.89 mg/kg after oral administration. All these characteristics make CR 2194 an important compound in the investigation of the biological effects of gastrin and a potential agent for diagnostic or therapeutic use.


Pharmacological assessment of Ca2+ dependence of endothelin-1-induced response in rat aorta. In rat aorta, endothelin-1 (ET-1) induced a slowly developing and sustained contraction in Ca(2+)-containing normal Krebs, nominally Ca(2+)-free Krebs, and Ca(2+)-free Krebs containing EGTA with a decreasing level of contraction. When ET-1-precontracted tissues were washed with Ca(2+)-free Krebs +50 microM EGTA, the tissues spontaneously and slowly relaxed. Readministration of Ca2+ during the early spontaneous relaxation phase caused a rapidly developing tonic contraction. When added during the late spontaneous relaxation phase, Ca2+ evoked slowly developing or, sometimes, biphasic contractions. In the presence of sarcoplasmic reticulum Ca(2+)-pump inhibitor, cyclopiazonic acid, the above biphasic contraction brought about by readministration of Ca2+ converted to a rapidly developing monophasic response. Thus, the first component of the biphasic contraction may be due to refilling of ET-1-sensitive Ca2+ store via the internal membrane Ca(2+)-pump. Furthermore, the ability of the phenylephrine-sensitive pool of internal Ca2+ to refill in the presence of 10(-8) M ET-1 suggests that the phenylephrine-sensitive pool differs from ET-1-sensitive pool and cannot be depleted by ET-1.


Relative agonist potencies of C2-substituted analogues of adenosine: evidence for adenosine A2B receptors in the guinea pig aorta. Nine C2-substituted adenosine analogues that are potent and selective for the A2-adenosine receptor were tested for their ability to induce relaxations of the guinea pig aorta. Compounds tested were 2-phenylethoxyadenosine (PEA), 2-phenylethoxy-5'-N-ethylcarboxamidoadenosine (PENECA), 2-cyclohexylethoxyadenosine (CEA), 2-fluorophenylethoxyadenosine (FPEA), 2-methoxyphenylethoxyadenosine (MPEA), 2-naphthylethoxyadenosine (NEA), 2-phenylaminoadenosine (CV-1808), 2-phenylethylaminoadenosine (PEAA) and 2-carboxyethylphenethylamino-5'-N-ethylcarboxamidoadenosine (CGS21680). The responses to these agents were compared to those of three standard adenosine receptor agonists, 5'-N-ethylcarboxamidoadenosine (NECA), N6-cyclohexyladenosine (CHA) and R-N6-phenylisopropyladenosine (R-PIA). The C2-ethoxyadenosine analogues were 30- to 140-fold less potent than NECA and the C2-amino-substituted analogues were 250 to 1000-fold less potent than NECA at inducing relaxations of the guinea pig aorta. All of the analogues were also less potent than the A1-selective agonist R-PIA. However, only responses to NECA were competitively antagonized by the non-selective adenosine receptor antagonist 8-phenyltheophylline (8-PT), pKB = 6.83 +/- 0.05. The results suggest that the C2-substituted analogues produce relaxations of the guinea pig aorta through a combination of actions at A2-adenosine receptors and at xanthine resistant sites. The lack of potency of these analogues at activating the xanthine sensitive A2-receptors in the guinea pig aorta suggests that these adenosine receptors may be of the A2b-subtype.


Displacement of in vivo binding of [3H]brofaromine to rat intestinal monoamine oxidase A by orally administered tyramine. The reversibility of the interaction of inhibitors with monoamine oxidase (MAO) is thought to provide a safety valve with respect to tyramine potentiation. We sought experimental evidence for this concept by studying the binding of orally administered [3H]brofaromine to the A-form of MAO in the rat ileum. Specific binding, defined by pretreatment with 10 mg/kg clorgyline p.o., amounted to 70-90% of total binding between 30 min and 6 h after administration of the radioligand. Brofaromine and clorgyline dose dependently displaced [3H]brofaromine with ED50 values of about 0.2 mg/kg when administered orally after the radioligand; so did orally administered tyramine in doses relevant for tyramine potentiation in the rat. It was also found that tyramine was relatively more effective in partially MAO-inhibited rats. The data suggest that the concept of reversibility functioning as a safety valve with respect to the potentially hazardous effects of tyramine ingestion is realistic.


Involvement of the cholinergic system in the effects of nefiracetam (DM-9384) on carbon monoxide (CO)-induced acute and delayed amnesia. The effects of N-(2,6-dimethylphenyl)-2-(2-oxo-1-pyrrolidinyl)-acetamide (DM-9384, nefiracetam), a cyclic derivative of GABA, were investigated in the carbon monoxide (CO)-induced amnesia model in mice using the passive avoidance task. Memory deficiency occurred when mice were exposed to CO before memory was completely consolidated after training (acute amnesia), at 7 days before training and 7 days after training (delayed amnesia). DM-9384 prolonged the step-down latency in mice with CO-induced amnesia. Scopolamine blocked the anti-amnesic effect of DM-9384 on delayed amnesia that had been induced by pre- or post-training exposure to CO. Bicuculline had a tendency to antagonize the anti-amnesic effect of DM-9384, but this tendency was not significant. Under these conditions, no significant change in the activity of choline acetyltransferase and glutamic acid decarboxylase was observed in the frontal cortex, striatum and hippocampus. These results suggest that DM-9384 potentiates cholinergic neuronal function and that it may modify acquisition and/or consolidation of memory.


Mechanical and ionic response of rat aorta to diazoxide. Diazoxide provoked concentration-dependent and endothelium-independent relaxations of the mechanical responses evoked by low concentrations of KCl. Glibenclamide, tolbutamide and tetraethylammonium shifted the concentration-response curve for diazoxide to the right. The drug also caused a dose-dependent stimulation of 86Rb outflow which was inhibited by glibenclamide and tolbutamide. Diazoxide (10(-4) and 10(-3) M) inhibited the contractions elicited by 10(-1) M K+ and provoked a concentration-dependent reduction in the contractile responses to Ca2+. Diazoxide also reduced the KCl (8 x 10(-2) M)-induced increase in 45Ca outflow. These data indicate that the vasorelaxant properties of diazoxide are probably related to an inhibition of Ca2+ entry into smooth muscle cells. The reduction in Ca2+ entry appears to result from K+ channel activation. At high concentrations, diazoxide also exhibited antagonistic actions on voltage-sensitive Ca2+ channels.


Hyperglucagonemia induced in mice by tryptamine: involvement of the peripheral 5-HT2 receptors. The effects of an indoleamine, tryptamine, on plasma glucagon levels were investigated in mice. Tryptamine induced dose-related increases in plasma glucagon levels. The hyperglucagonemia effects of tryptamine were completely antagonized by methysergide and ketanserin which have a high affinity to 5-HT2 receptors. In addition, the peripheral 5-HT2 receptor antagonist, xylamidine, also strongly inhibited tryptamine-induced hyperglucagonemia. Our results indicate that the peripheral 5-HT2 receptors mediate the increase in plasma glucagon levels induced by tryptamine and that these receptors may have a role in the control of glucagon secretion.


Contractile activity of vasotocin, oxytocin, and vasopressin on mammalian prostate. The neurohypophyseal peptides arginine vasotocin, oxytocin and arginine vasopressin contracted guinea pig, rat, canine and human prostates with potencies and efficacies that were comparable to those of noradrenaline and methacholine. All three neuropeptides raised prostatic tone and elicited contractions at 10(-9) or 10(-8) M, with an order of efficacy: arginine vasotocin greater than oxytocin greater than arginine vasopressin. The findings suggest a physiological role for these peptides in prostatic smooth muscle contraction and possibly also in other aspects of male reproductive function.


PF-5901 inhibits gastrointestinal platelet-activating factor synthesis in vivo. PF-5901, a novel anti-inflammatory compound, has previously been shown to inhibit the synthesis of platelet-activating factor (PAF) by peritoneal mast cells in vitro. The purpose of the present study was to assess the effects of this compound on PAF synthesis in vivo using two animal models which are characterized by elevated levels of gastrointestinal PAF synthesis. In the first study, rats were infected with Nippostrongylus brasiliensis and killed 17 days later. Groups of infected rats were given PF-5901 at doses ranging from 0.1 to 100 mg/kg, and 12 h later the effects on jejunal PAF synthesis were determined. PF-5901 reduced PAF synthesis in a concentration-dependent manner, with inhibition reaching 95% at the highest dose tested. In the second study, groups of rats were orally pretreated with PF-5901 (100 mg/kg) or vehicle 3 or 12 h prior to induction of endotoxic shock by i.v. administration of lipopolysaccharide from Salmonella typhosa. PAF synthesis in various regions of the gastrointestinal tract was assessed 15 min later. PF-5901 significantly reduced the hemoconcentration, but not the hypotension associated with endotoxic shock. Moreover, this compound significantly inhibited PAF synthesis in all tissues studied when a 12 h pretreatment time was used, and in all tissues except the duodenum when a 3 h pretreatment time was used. These studies demonstrate that PF-5901 inhibits gastrointestinal PAF synthesis in two in vivo models. This compound may therefore be a useful pharmacological tool for future studies on the role of PAF in inflammatory processes, and the effects of PF-5901 on PAF synthesis may contribute to its anti-inflammatory properties.


Effects of verapamil on [3H]acetylcholine release in the striatum of spontaneously hypertensive rats. In the present study, we describe the effects of Ca2+ channel antagonist (verapamil) on [3H]acetylcholine (ACh) release in the central nervous system of spontaneously hypertensive rats (SHR). The electrically stimulated release of [3H]ACh from striatal slices was not different between SHR and normotensive Wistar Kyoto (WKY) rats. Verapamil inhibited electrically stimulated [3H]ACh release in a dose-related fashion. The inhibitory effect of verapamil was significantly greater in SHR than in WKY rats. These results suggest that the Ca2+ sensitivity of central cholinergic neurons might be enhanced in SHR, which could attribute, at least partially, to the pathogenesis of hypertension.


8-Methoxypsoralen blocks ATP-sensitive potassium channels and stimulates insulin release. 8-Methoxypsoralen (8-MOP) stimulated insulin release from HIT-T15 B-cells and inhibited the diazoxide-induced and sulfonylurea-sensitive 86Rb+ efflux from these cells. These results indicate that 8-MOP affects ATP-sensitive K+ channel activity. Patch-clamp experiments confirmed this view.


Hippocampal lesions induced by microinjection of the nitric oxide donor nitroprusside. Unilateral intrahippocampal microinjection of the nitric oxide (NO) donor, sodium nitroprusside (33 nmol in 1 microliter), in the rat resulted in a marked degeneration of the ipsilateral hippocampal formation characterised by a virtual absence of pyramidal cells and dentate granule cells. Damage was not observed contralateral to the injection site. Quinolinic acid (100 nmol in 1 microliter) also produced neuronal damage within the ipsilateral hippocampus although the lesion was considerably smaller and more discrete than that caused by sodium nitroprusside. Injection of equimolar amounts of potassium ferricyanide failed to mimic the neurotoxic effects of sodium nitroprusside suggesting that NO is responsible specifically for the neuronal damage observed.


Activation of metabotropic receptors has a neuroprotective effect in a rodent model of focal ischaemia. The role of the glutamate 'metabotropic' receptor was investigated in an experimental model of focal ischaemia-induced neurodegeneration. The metabotropic agonist, trans-1-amino cyclopentane-1,3-dicarboxylic acid (t-ACPD, 20 mg/kg i.p.), was administered to mice immediately after middle cerebral artery occlusion (MCAO), which causes cerebral infarct. Seven days after MCAO, the mean infarct volume value of the t-ACPD-treated group (mean +/- S.E. = 4.57 +/- 0.73 mm3) was significantly reduced, by 34.3%, compared to the vehicle-treated group (mean +/- S.E. = 6.95 +/- 0.59 mm3, P less than 0.01). This suggests that metabotropic receptor activation in the adult brain reduces excitotoxicity.


Competitive NMDA receptor antagonists attenuate phencyclidine-induced excitations of A10 dopamine neurons. The binding of phencyclidine (PCP) within the channel gated by the NMDA-receptor complex can be positively or negatively modulated by compounds which facilitate or prevent the interaction of glutamate to the NMDA recognition site. In the present study extracellular recordings were used to evaluate the possibility that the negative modulation of NMDA channel function by the competitive NMDA antagonists (+/-)-CPP (3-((+/-)2-carboxypiperazin-4- yl)propyl-1-phosphonate) and CGS 19755 (cis-4-phosphonomethyl-2-piperidine carboxylate) could affect the response of A10 dopamine neurons to PCP. Pretreatment with 40 mg/kg of (+/-)-CPP or CGS 19755 completely blocked the low-dose excitatory effects of PCP, whereas 10 mg/kg of CGS 19755 produced only a partial blockade. However, neither CGS 19755 or (+/-)-CPP affected the amount of attenuation of A10 firing occurring with large doses of PCP. (+/-)-CPP and CGS 19755 pretreatment also failed to alter the morphine-induced stimulation of dopamine activity. These findings not only provide further evidence that the low-dose PCP-induced activation of A10 neurons is mediated through the NMDA-ion channel complex, but suggest that some physiological or behavioral effects evoked by PCP might be prevented by treatment with competitive NMDA receptor blockers.


Effects of BY-1949 on evoked responses of cortical blood flow and Meynert neurons. The effects of BY-1949, a novel dibenzoxazepine derivative, injected into the cerebral ventricle on noxious stimulus-induced responses of regional blood flow in the cortex and neuronal activity in the nucleus basalis of Meynert were studied in male rats. These induced responses were markedly enhanced by administration of BY-1949 (16.4 +/- 0.5 ng/100 g, S.E.M.). These data indicate that BY-1949 acts on the central nervous system to modulate the response to a noxious stimulus of regional cerebral blood flow and neuronal activity in the nucleus basalis of Meynert.


Dopamine and melatonin in the nucleus accumbens may be implicated in the mode of action of antidepressant drugs. The effect of chronic systemic treatment (once a day for 2-3 weeks) with different antidepressant drugs (desipramine, mianserin, fluvoxamine 15 mg/kg per day i.p. or s.c.) on the behavioral responses elicited by intra-accumbens injection of graded doses (1-10,000 ng) of apomorphine and two doses (10 and 100 ng) of melatonin was investigated in rats. Treatment with antidepressant drugs consistently facilitated apomorphine-induced hypermotility, but differentially influenced apomorphine-induced hypomotility. The drugs completely antagonized melatonin-induced behavioral changes (hypomotility and increased duration of sniffing). The data suggest that chronic treatment with antidepressant drugs results in postsynaptic dopamine receptor supersensitivity in the nucleus accumbens, a terminal area of the mesolimbic dopaminergic system. It is postulated that the development of supersensitivity may be mediated by the inhibition of melatonin-induced effects observed after acute and chronic treatment with these antidepressants. The present findings may be relevant for the mode of therapeutic action of antidepressant drugs.


Involvement of a phosphoramidon-sensitive endopeptidase in the processing of big endothelin-1 in the guinea-pig. In anaesthetized and ventilated guinea-pigs, i.v. injection of 1 nmol/kg big endothelin-1 (big ET-1) did not evoke significant changes in pulmonary inflation pressure (PIP) and mean arterial blood pressure (MBP), whereas injection of the same dose of endothelin-1 (ET-1) induced marked and rapid bronchoconstrictor and pressor responses. Administered at the dose of 10 nmol/kg, big ET-1 provoked significant increases in PIP and MBP, which developed slowly and were long-lasting as compared to those evoked by ET-1. When big ET-1 was incubated for 45 min at 37 degrees C with alpha-chymotrypsin (2 mU/nmol) or pepsin (1 microgram/nmol) and then injected into guinea-pigs at the dose of 1 nmol/kg, marked bronchoconstrictor and pressor responses were observed, with kinetics similar to those noted after administration of the same dose of ET-1. The magnitude of the alpha-chymotrypsin- or pepsin-treated big ET-1 responses was similar to that induced by ET-1, incubated or not with the enzymes. Injected i.v. at the dose of 5 mg/kg, 5 min before the challenge, phosphoramidon almost totally inhibited the bronchoconstrictor and pressor responses induced by 10 nmol/kg big ET-1, whereas thiorphan (5 mg/kg) partially reduced the increase in PIP and exerted a minimal effect on the changes in MBP. Administered at the dose of 20 mg/kg per os, 1 h before i.v. administration of 10 nmol/kg big ET-1, enalapril maleate and captopril did not significantly alter the bronchoconstriction and the hypertensive response evoked by the peptide.(ABSTRACT TRUNCATED AT 250 WORDS)


Endothelin-1-induced prostaglandin E2 production: modulation of contractile response to endothelin-1 in porcine coronary artery. Endothelin-1 (ET-1, 1 nM) increased the release of prostaglandin E2 (PGE2) in endothelium-denuded smooth muscle strips of porcine coronary arteries. Indomethacin enhanced the amplitude of contraction induced by ET-1 and inhibited the stimulated release of PGE2. PGE2 (0.1-100 nM) attenuated the amplitude of contraction induced by 1 nM ET-1. These results suggest that in the smooth muscle of porcine coronary arteries, ET-1 increased the synthesis of PGE2, which functionally antagonizes the direct vasoconstrictor actions of ET-1.


Acetyl-L-carnitine releases dopamine in rat corpus striatum: an in vivo microdialysis study. The effect of acetyl-L-carnitine, a compound reported to be beneficial for senile patients, on the release of dopamine (DA) from the striatum was studied by using in vivo brain dialysis in anesthetized rats coupled with HPLC-electrochemical detection. Striatal infusion of acetyl-L-carnitine increased the efflux of DA with no apparent changes in efflux of DA metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and 4-hydroxy-3-methoxyphenylacetic acid (HVA). The DA-releasing effect of acetyl-L-carnitine was concentration- and Ca(2+)-dependent, and was abolished by omega-conotoxin fraction GVIA and tetrodotoxin, inhibitors of the voltage-dependent Ca2+ and Na+ channels, respectively. Nomifensine, an inhibitor of DA reuptake did not alter the DA-releasing property of acetyl-L-carnitine. DA released from the striatum by acetyl-L-carnitine was decreased by reserpine pretreatment whereas the d-amphetamine-evoked DA outflow was not affected. In contrast to acetyl-L-carnitine, d-amphetamine reduced the extracellular concentrations of DOPAC and HVA. We conclude from the present data that acetyl-L-carnitine evokes DA release from the vesicular pools of the nigrostriatal dopaminergic neurons by a Ca(2+)-dependent, exocytotic process.


Discriminative stimulus properties of the muscarinic receptor agonists Lu 26-046 and O-Me-THPO in rats: evidence for involvement of different muscarinic receptor subtypes. The discriminative cues induced by the muscarinic receptor agonists Lu 26-046 ((-)-7-methyl-3(2-propynyloxy)-4,5,6,7-tetrahydroisothiazolo [4,5-c]pyridine ) and O-Me-THPO (3-methoxy-4,5,6,7-tetrahydroisoxazolo[4,5-c]pyridine) were investigated. The results were compared with those obtained for the binding profiles of these agonists at central muscarinic receptors and with results concerning their functional effects at peripheral muscarinic receptors in vitro. Lu 26-046 had preferential affinity for M1 versus M2 receptors (Ki index [3H]quinuclidinyl benzilate ([3H]QNB/[3H]pirenzepine 4.2) and had partial agonistic activity at M1 and M2 receptors in rat superior cervical ganglion and guinea pig left atrim, respectively. A weak antagonistic effect at M3 receptors in guinea pig ileum was observed. O-Me-THPO had non-selective agonistic effects at peripheral M1, M2 and M3 receptors and had a slight preference for central M2 receptors in binding experiments (M2/M1 index 0.31). Lu 26-046 dose dependently substituted for Lu 26-046 and partially substituted for O-Me-THPO in rats trained to discriminate Lu 26-046 and O-Me-THPO from saline, respectively. The (+)-enantiomer of Lu 26-046, Lu 26-047, had weak partial M1 agonistic activity and M2/M3 antagonistic effects at peripheral receptors. Lu 26-047 also had a high M2/M1 index (9.3) in binding experiments. Lu 26-047 substituted for Lu 26-046, but preferentially inhibited the effect of O-Me-THPO. Pilocarpine had a preferential effect in Lu 26-046-trained rats, while oxotremorine and arecoline had preferential effects in O-Me-THPO-trained rats. Large increases in latency times or a disruption of responding was generally observed. These compounds were full agonists at peripheral M1, M2 and M3 receptors. The muscarinic receptor antagonist scopolamine antagonized the effect of O-Me-THPO and partially inhibited the effect of Lu 26-046. Scopolamine partially substituted for Lu 26-046. The quaternary muscarinic receptor agonist N-methyl atropine had no effect, indicating that the cues are mediated by central muscarinic receptors. It is suggested that the discriminative cues of Lu 26-046 and O-Me-THPO are preferentially mediated by central M1 (partial) and M2 receptor stimulation, respectively. The role of central M3 receptors is not known.


Decreased activity of rat A10 dopamine neurons following withdrawal from repeated cocaine. Electrophysiological techniques were used to determine the basal activity of A10 dopamine (DA) neurons in the rat ventral tegmental area after a 10-14 day withdrawal from repeated cocaine treatment (10.0 mg/kg i.p. twice daily for 14 days). The number of spontaneously active A10 DA cells was significantly decreased (42%) in the cocaine-treated rats. This decreased activity may underlie the diminished basal levels of synaptic DA within the nucleus accumbens previously reported in cocaine-withdrawn rats and may account for anhedonia, anergia and cocaine craving reported by withdrawn cocaine addicts.


Dopamine transport: pharmacological distinction between the synaptic membrane and the vesicular transporter in rat striatum. The pharmacological properties of the monoamine transporters in the synaptic vesicles and of the dopamine transporters in the synaptic plasma membrane were compared. Tetrabenazine, an inhibitor of the vesicular transporter did not block ligand binding to the plasma membrane transporter. Various potent cocaine analogues and other compounds active at the plasma membrane transporter did not block ligand binding to the vesicular transporter. These data indicate pharmacological differences between the vesicular and synaptic membrane transporters.


The effect of calcium channel modulators on blood pressure and pressor responses to noradrenaline in the guinea-pig. Selective L-type voltage-operated calcium channel (VOCC) antagonists nicardipine, diltiazem and verapamil all produced pronounced falls in mean arterial blood pressure (MAP) in the anaesthetized artificially ventilated guinea-pig. This fall in MAP was associated with a significant reduction of the pressor response induced by i.v. noradrenaline. omega-Conotoxin GVIA (CgTx) a preferential N-type VOCC antagonist produced a significant time-dependent fall in MAP, similar in magnitude to the L-type VOCC antagonist, but did not affect the noradrenaline-induced pressor responses.


Preferential small arteriolar vasodilatation by the potassium channel opener, BRL 38227, in conscious spontaneously hypertensive rats. The microvascular effects of the potassium channel opener, BRL 38227 (the active enantiomer of cromakalim), were investigated in conscious spontaneously hypertensive rats (SHR) provided with a chronic dorsal microcirculatory chamber. BRL 38227 decreased blood pressure and increased heart rate in a dose-dependent manner. It dilated arterioles of different sizes, with the most pronounced effect on the smallest precapillary arterioles. Venule diameters were not significantly affected by BRL 38227. The data show a preferential sensitivity of small precapillary arterioles for potassium channel opening.


Neuroprotective effects of NG-nitro-L-arginine in focal stroke in the 7-day old rat. Recent evidence in primary neuronal cell culture implicates nitric oxide (NO) as a mediator of glutamatergic neurotoxicity acting via N-methyl-D-aspartate (NMDA) receptors. We find that administration of the potent nitric oxide synthetase (NOS) inhibitor NG-nitro-L-arginine (NO-Arg) at 50 mg/kg to 100 mg/kg i.p. to 6-day old Sprague-Dawley rat pups results in prompt and long-lasting in vivo inhibition of NOS. Fifteen hours after administration, NO-Arg produces essentially complete neuroprotection against hypoxic-ischemic in a standard (Rice-Vanucci) model. These results support the hypothesis that NO may play a key mediatory role in brain damage attending focal ischemic stroke.


Effect of Ca2+ modulators on acetylcholine-induced phasic and tonic contractions and A23187-induced contractions in ileal longitudinal muscle and IP3 production. The influence of different sources of Ca2+ on the Emax and ED50 values of acetylcholine (ACh)-induced phasic and tonic contractions and on A23187-induced contractions was studied using different extracellular Ca2+ concentrations ([Ca2+]o) and the Ca2+ modulators TMB-8 and D600. IP3 production induced by both stimulants was also studied. The results are compatible with: (a) the mobilization of Ca2+ from an intracellular source as a primary event in the phasic response. (b) The primary involvement of a D600-sensitive inward Ca2+ current in the ACh-tonic response. (c) An inward D600-sensitive Ca2+ current associated with the ionophore transported ion. (d) The involvement of an IP3 independent, TMB-8 sensitive mechanism of Ca2+ mobilization involved in the A23187-induced responses.


Characterization of BIBS 39 and BIBS 222: two new nonpeptide angiotensin II receptor antagonists. Two new nonpeptide angiotensin II (AII) receptor antagonists, 4'-[(2-n-butyl-6-cyclohexylaminocarbonylamino-benzimidazole-1-yl)- methyl ] biphenyl-2-carboxylic acid (BIBS 39) and 2-n-butyl-1-[4-(6-carboxy-2,5-dichlorobenzoylamino)-benzyl]-6-N- (methylaminocarbonyl)-n-pentylamino-benzimidazole (BIBS 222) were characterized in radioligand binding assays, and in vitro and in vivo experiments. BIBS 39 displaced [125I] AII from its specific binding sites with a K(i) value of 29 +/- 7 nM for the AII subtype I (AT1) receptor and a K(i) value of 480 +/- 110 nM for the AII subtype 2 (AT2) receptor. BIBS 222 showed a K(i) value of 20 +/- 7 nM for the AT1 subtype and a K(i) value of 730 +/- 170 nM for the AT2 subtype. Thus BIBS 39 was 17 times more selective for the AT1 subtype and BIBS 222 37 times. Both compounds were specific for AII receptors as they did not show high affinity for other receptors. BIBS 39 shifted the AII concentration-contractile response curves in isolated rabbit aorta to the right in a parallel fashion. A pA2 value of 8.14 +/- 0.08 and a slope of 1.06 +/- 0.07 were calculated. BIBS 222 caused nonparallel shifts to the right and reduced the maximal response induced by AII by about 25%. A KB value of 9.01 (+/- 3.22) x 10(-8) M was determined. At 10(-5) M, neither compounds altered the contractile responses to noradrenaline and KCl. In pithed rats, BIBS 39 dose dependently shifted the dose-response curve of AII to the right without affecting the maximal response.(ABSTRACT TRUNCATED AT 250 WORDS)


Modification of chlordiazepoxide's behavioural and neurochemical effects by handling and plus-maze experience. The purpose of the present experiment was to determine how a rat's prior history (of repeated gentle handling and/or of the elevated plus-maze apparatus) modified the behavioural and neurochemical response to chlordiazepoxide. In handled animals one previous exposure to the plus-maze rendered the rats insensitive to the anxiolytic effects of chlordiazepoxide in this test. This phenomenon of 'one-trial tolerance' was not seen in unhandled rats and thus both prior handling and prior maze experience were necessary to abolish the behavioural response to chlordiazepoxide. The effects of chlordiazepoxide on K(+)-evoked [14C]GABA (gamma-aminobutyric acid) release were also modified by the rat's past history. The drug-induced reduction of GABA release in the cortex was abolished by prior plus-maze experience; whereas handling modified chlordiazepoxide's effects on GABA release in the hippocampus (the drug decreased release in unhandled rats and increased release in those given repeated gentle handling). Thus an anxiolytic response to chlordiazepoxide in the plus-maze was accompanied by reduced GABA release in both cortex and hippocampus. The 5-HT system (5-hydroxytryptamine) also proved sensitive to the rats' past history. The effects of chlordiazepoxide on K(+)-evoked [3H]5-HT release from the hippocampus depended on both prior handling and plus-maze experience and could be predicted from the undrugged level of evoked release; when this was low, chlordiazepoxide increased it, when it was high, chlordiazepoxide reduced it. These results raise the possibility that the beneficial effects of a benzodiazepine may depend on the baseline condition of the animals.


Quinacainol, a new antiarrhythmic with class I antiarrhythmic actions in the rat. The antiarrhythmic and electrophysiological actions of quinacainol, a new Class I antiarrhythmic, were assessed in rats. Electrophysiological actions of quinacainol were assessed in vivo in terms of drug-induced changes in ECG, responses to left ventricular electrical stimulation, and changes in epicardial intracellular potentials to precisely characterize the electrophysiological effects of this putative subclass Ic antiarrhythmic compound. Antiarrhythmic actions were assessed in conscious rats subjected to occlusion of the LAD coronary artery. Antiarrhythmic actions occurred with 2.0 and 4.0 mg/kg, whereas 8.0 mg/kg was pro-arrhythmic. At doses of 0.5 mg/kg and above quinacainol increased threshold currents for capture and for ventricular fibrillation. Doses of 2.0 mg/kg and above increased ventricular refractoriness. From 1.0 to 8.0 mg/kg, quinacainol reduced dV/dtmax of phase 0 of epicardial action potentials but only 8.0 mg/kg increased action potential duration. The Q-T interval was also increased with the highest dose. Quinacainol dose-relatedly increased P-R interval whereas QRS did not change. Thus the Class I electrophysiological properties of quinacainol over the dose range tested did not fit accurately into a single subclass of the various subclasses of Class I. However, the Class Ic actions seen with 2.0 and 4.0 mg/kg were associated with antiarrhythmic actions.


Effects of the putative 5-HT1A receptor antagonist NAN-190 on free feeding and on feeding induced by the 5-HT1A receptor agonist 8-OH-DPAT in the rat. The effects of the putative 5-HT1A receptor antagonist NAN-190 on feeding and spontaneous locomotor activity in rats were examined. The drug elicited a robust, dose-dependent (0.01-10 mg/kg) increase in food consumption in free feeding animals. Microstructural analysis of feeding induced by NAN-190 (3 mg/kg) revealed that the drug increased the duration of feeding and number of feeding bouts but decreased the feeding rate. The increase in feeding induced by 3 mg/kg of NAN-190 was not apparent until 2-4 h after injection. This prolonged latency to onset of the feeding response appeared to be due to response competition. Thus, a 'neuroleptic-like' action of the drug on spontaneous motor activity was observed during the the initial 2 h following injection. A dopamine receptor antagonist action of NAN-190 was also indicated by the results of studies in which the drug was observed to block oral stereotypy induced by the dopamine receptor agonist apomorphine. In interaction studies, NAN-190 (0.1 and 10 mg/kg) failed to block the feeding response induced by the prototypical 5-HT1A receptor agonist 8-OH-DPAT (0.0625 and 1.0 mg/kg) and indeed, appeared to have an additive effect with 8-OH-DPAT on consummatory behaviour. These data suggest that NAN-190 may act as a partial agonist rather than an antagonist at the 5-HT1A receptor and also provide the first evidence that the drug has dopamine receptor antagonist properties in vivo.


Effects of two different reversible monoamine oxidase-A inhibitors on nociceptive thresholds in the rat. The acute effect of two different reversible inhibitors of monoamine oxidase-A on nociceptive thresholds was evaluated in the rat by the tail-flick and hot-plate tests. CGP 11305-A, a monoamine oxidase-A inhibitor that also blocks serotonin reuptake, elicited an increase of latency in the tail-flick and the hot-plate test. Ineffective doses of CGP 11305-A increased nociceptive thresholds when administered in combination with other serotoninergic agents, i.e. chlorimipramine or 5-hydroxytryptophan, at doses that were ineffective alone. CGP 22364-A, a pure inhibitor of monoamine oxidase-A, increased latency only in the hot-plate test. Both compounds decreased spontaneous locomotor activity at the doses effective in the hot-plate test, suggesting that the responses observed in this test are not related to a pure effect on nociceptive thresholds. The data suggest that the increase in serotonin availability induced by monoamine oxidase-A inhibition alone is not sufficient to affect nociceptive thresholds.


13,14-Dihydro-prostaglandin E1 decreases low-density lipoprotein influx into rabbit aorta. The effect of 4-week daily administration of 13,14-dihydro-prostaglandin E1 (13,14-DH-PGE1; 2 micrograms/kg) on LDL influx into the rabbit aortic wall was examined versus the effect of the same dose of PGE1 and sham-treatment in 108 male animals fed an 1% cholesterol supplemented diet. Treatment was started after de-endothelialization of the abdominal aorta with a Fogarty catheter. After the 1-month treatment period the animals were injected with 10 microCi 125I-low-density lipoprotein (LDL; 0.5 mg protein/ml). Uptake of the radiolabelled LDL was measured in morphologically verified endothelialized, re-endothelialized and de-endothelialized abdominal aortic segments. The LDL influx into the aorta was significantly (P less than 0.001) lower in 13,14-DH-PGE1- and PGE1-treated rabbits than in the controls. This reduction was most pronounced in re- and de-endothelialized segments. No significant difference between effect of PGE1 and of its biologically active derivative, 13,14-DH-PGE1, on arterial LDL entry was found. These data demonstrate a comparable beneficial effect of 13,14-DH-PGE1 and PGE1 on vascular wall lipid metabolism by decreasing LDL entry into the aortic wall in vivo.


AS-5370 potently antagonizes 5-HT3 receptor-mediated responses on NG108-15 cells and on the rat vagus. The action of a novel 5-HT3 receptor antagonist, AS-5370, has been studied on two electrophysiological models for 5-HT3 receptors: whole-cell patch-clamp recordings from mouse neuroblastoma-rat glioma (NG108-15) cells and grease-gap recordings from rat isolated vagus nerve. The 5-hydroxytryptamine (5-HT)-induced fast inward current of voltage-clamped NG108-15 cells was antagonized by 1 nM AS-5370 in an insurmountable manner. On the rat vagus, AS-5370 reduced the maximum depolarizing response to 5-HT in a concentration-dependent manner. The IC50 for AS-5370 on the rat vagus was 0.3-1.0 nM. The EC50 for 5-HT on the rat vagus was not appreciably affected by AS-5370. On the rat vagus, the (R) enantiomer of AS-5370 was about 30 times more potent than the (S) enantiomer. The antagonist action of AS-5370 on these two cell types was compared with that of (+)-tubocurarine. Unlike tubocurarine, the effect of AS-5370 on NG108-15 cells was not readily reversed during wash. On the rat vagus, tubocurarine antagonized in a competitive manner with an IC50 of 0.3-1.0 microM (pKb = 7.2). It is concluded that AS-5370 is a potent 5-HT3 receptor antagonist on both NG108-15 cells and the rat vagus, but it does not act in a competitive manner.


Direct and indirect effects of angiotensin II on venous tone in conscious rats. The direct and indirect effects of angiotensin II (ANGII) on mean arterial pressure (MAP) and mean circulatory filling pressure (MCFP), an index of body venous tone, were investigated in conscious rats. Dose-response curves of ANGII were constructed in control rats (Group I), rats pretreated with saralasin (competitive ANGII antagonist, Group II), with guanethidine (inhibitor of sympathetic postganglionic neurons. Group III), or the ganglionic blocker hexamethonium (Group IV) and rats given unilateral right adrenalectomy two days prior to the study (Group V). The infusion of single doses of ANGII in control, adrenalectomized, guanethidine-treated and hexamethonium-treated rats dose dependently increased MAP to similar maxima; ED50 value was increased by adrenalectomy but unaffected by guanethidine nor hexamethonium. The pressor effects of ANGII was almost completely abolished by saralasin. ANGII dose dependently increased MCFP in control rats. In hexamethonium-treated rats, ANGII also dose relatedly increased MCFP which reached similar maximum as that in control rats, but the ED50 value was reduced. Saralasin almost completely abolished the MCFP response. Both guanethidine and adrenalectomy reduced maximum MCFP response to ANGII, but neither altered the ED50 value. Our results show that the sympathetic nervous system contributed greater to the MCFP than MAP effects of ANGII. Both direct and indirect effects of ANGII are mediated via the activation of ANGII receptors that are susceptible to blockade by saralasin.


Losartan potassium, a nonpeptide antagonist of angiotensin II, chronically administered p.o. does not readily cross the blood-brain barrier. Recently several novel nonpeptide antagonists of angiotensin II (Ang II) have been identified. One of these, losartan potassium (formerly DuP 753) was developed as an orally active and highly selective antagonist for Ang II. As it is inhibited by sulfhydryl agents, it is specific for the AT1 receptor subtype. Since Ang II has both central and peripheral effects, we investigated whether losartan, given p.o. chronically, crosses the blood-brain barrier. The effects of chronic administration of losartan orally (p.o.) at 3 mg/kg per day for three days on the dipsogenic and pressor responses to a pre-established dose of Ang II i.v.t. (50 ng) were studied. Three series of experiments were carried out using conscious normotensive Sprague-Dawley rats. The rats were injected with Ang II intraventricularly (i.v.t.) before and after treatment of losartan p.o. and blood pressure and drinking responses measured. The experiments established that 3 mg/kg losartan p.o. for 3 days antagonized pressor effects of Ang II intravenously (i.v.), but did not antagonize the pressor or drinking effects of Ang II i.v.t. Daily water intake significantly increased with chronic losartan p.o.. Since chronic administration of losartan p.o. was able to block the effects of Ang II i.v. but had no effect on Ang II i.v.t. we conclude that losartan potassium does not readily cross the blood-brain barrier using this dose regimen.


Effect of SCH 37224 on anti-IgE-induced formation of sulfidopeptide leukotrienes in human lung parenchyma. SCH 37224, 1-(1,2-dihydro-4-hydroxy-2-oxo-1-phenyl-1,8-naphthyridin-3-yl) pyrrolidinium, is a structurally novel compound that had been shown to inhibit leukotriene D4 formation in guinea pig lung in vitro. We tested whether SCH 37224 is able to inhibit both the formation of eicosanoids from human lung parenchyma in vitro and the binding of sulfidopeptide leukotrienes to membranes of lung parenchyma and bronchi. SCH 37224, at a concentration of 30 and 100 microM, was able to inhibit antigen-induced formation of sulfidopeptide leukotrienes, measured as leukotriene E4, while it did not significantly affect the formation of prostaglandin D2. At concentrations up to 100 microM, it did not affect either the binding of [3H]leukotriene C4 to membranes of human lung parenchyma or human bronchi, or the binding of [3H]leukotriene D4 to the parenchyma. In conclusion, SCH 37224 is a selective inhibitor of leukotriene formation in human lung in vitro, which might be of potential therapeutic interest in the treatment of asthma.


8-OH-DPAT-induced inhibition of renal sympathetic nerve activity and serotonin neuronal firing. The effects of the 5-HT1A receptor 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) administered i.v. on medullary 5-HT neuronal firing and renal sympathetic nerve activity were determined in spontaneously breathing anaesthetized cats. Low doses of 8-OH-DPAT (1-3 micrograms/kg) caused a similar reduction in 5-HT neuronal firing and renal nerve activity while high doses (10-30 micrograms/kg) completely inhibited neuronal firing but caused only 80% inhibition of renal nerve activity. These data are discussed in relationship to the mechanism of sympatholytic action of 8-OH-DPAT and the serotonergic regulation of sympathetic nerve activity.


Inhibition of locus coeruleus neurons by the phencyclidine analog, N-[1-(2-benzo(b)thiophenyl)cyclohexyl]piperidine: evidence for potent indirect adrenoceptor agonist properties. The effects of the phencyclidine derivative, N-[1-(2-benzo(b)thiophenyl)cyclohexyl]piperidine (BTCP), on the electrical activity of noradrenaline (NA) neurons of the locus coeruleus (LC) were studied in halothane-anesthetized rats. Systemic administration of BTCP potently inhibited LC neurons (ID50 of 1.1 +/- 0.1 mg/kg i.v.). This effect was mimicked by local microejection of BTCP into the LC. Both the systemic and local effects of BTCP were blocked by alpha 2-adrenoceptor antagonists and prevented by prior depletion of catecholamines with reserpine. These and other data suggest that BTCP behaves as a potent indirect NA agonist (i.e. via NA re-uptake and/or release systems).


Activation of the glutamate metabotropic receptor protects retina against N-methyl-D-aspartate toxicity. Intraocular pretreatment with the specific metabotropic glutamate receptor agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) in the adult rat reduced the excitotoxic effects induced in the retina by a single intraocular injection of N-methyl-D-aspartate (NMDA). Damage was estimated by assessing NMDA-induced loss of retinal choline acetyltransferase (ChAT) activity. The interaction between metabotropic and ionotropic glutamate receptors may, therefore, be considered an important target for in vivo pharmacological neuroprotection.


Effect of SCH 23390 and quinpirole on novelty-induced grooming behaviour in spontaneously hypertensive rats and Wistar-Kyoto rats. Grooming behaviour induced by exposure to a novel environment was studied in spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). The dopamine D1 receptor antagonist, SCH 23390, and the dopamine D2 receptor agonist, quinpirole, were used to study brain dopamine systems in these rat strains, via their effects on grooming behaviour. The total grooming behaviour displayed in a 50-min observation period was significantly lower in SHR than in WKY. Except for the paw licking component no differences between the two strains were observed in the separate behavioural elements of grooming behaviour. SCH 23390 and quinpirole were found to suppress novelty-induced grooming behaviour of both strains. In SHR, grooming behaviour was less suppressed by SCH 23390, whereas the suppression by quinpirole was more pronounced than in WKY. These results indicate that there are alterations in central dopamine systems in SHR, probably involving changes both in dopamine D1 and D2 receptor mechanisms in the brain.


Dopamine receptor occupancy in vivo: behavioral correlates using NNC-112, NNC-687 and NNC-756, new selective dopamine D1 receptor antagonists. The ability of dopamine D2, mixed D1/D2 and selective D1 receptor antagonists, including NNC-112, NNC-687, NNC-756, to inhibit the in vivo binding of [3H]SCH 23390 or [3H]raclopride to dopamine receptors was studied in mice and rats. Furthermore, the dopamine-antagonistic effects of these drugs were also studied in various behavioral models. Significant levels of in vivo receptor blockade were required for antagonism of typical dopamine agonist-mediated behaviors. However, fewer D1 than D2 receptors had to be blocked to produce similar antagonistic effects. Thus, there may be a greater receptor reserve for D2 receptors than for D1 receptors.


NNC-112, NNC-687 and NNC-756, new selective and highly potent dopamine D1 receptor antagonists. The neurochemical properties of three novel benzazepine derivatives NNC-112, NNC-687 and NNC-756 were assessed. These compounds inhibited dopamine D1 receptor binding in vitro with low nanomolar to picomolar dissociation constants whereas those for the D2 receptor were in the micromolar range. Contrary to classical neuroleptics, but similar to the atypical neuroleptics, clozapine and fluperlapine, NNC-112, NNC-687 and NNC-756 were relatively more potent in inhibiting dopamine-stimulated adenylyl cyclase than [3H]SCH 23390 binding. Both NNC-112 and NNC-756 had high affinity for the 5-HT2 receptor whereas NNC-687 had low affinity for this receptor. The affinity for other receptors or neurotransmitter transporters was very low. In vivo, the dopamine D1 receptor selective profile of NNC-112, NNC-687 and NNC-756 was evident from the potent inhibition of D1 receptor binding whereas no effect on D2 receptor binding was apparent. In addition, the compounds blocked D1 receptor-mediated rotation in unilaterally 6-hydroxydopamine-lesioned rats, but had no effect on D2-induced rotation. Thus, NNC-112, NNC-687 and NNC-756 are potent and selective dopamine D1 receptor antagonists that may be useful in the treatment of schizophrenia.


Amitriptyline, desipramine, cyproheptadine and carbamazepine, in concentrations used therapeutically, reduce kainate- and N-methyl-D-aspartate-induced intracellular Ca2+ levels in neuronal culture. The glutamate receptor agonists, kainate and N-methyl-D-aspartate (NMDA) result in the elevation of intracellular calcium levels ([Ca2+]i) in primary cultures of cerebellar granule neurons. Several tricyclic antidepressants (TCAs), amitriptyline (0.5-1 microM), desipramine (1 microM) and doxepine (1 microM) partially prevent this elevation induced by both of these excitatory amino acids (EAAs), but not elevations of [Ca2+]i induced by another EAA, quisqualate. Evidence suggests that this EAA-tricyclic interaction may involve voltage-dependent Ca2+ channels since amitriptyline also partially blocks the elevation of [Ca2+]i induced by membrane depolarization with 40 mM KCl. However, the blockade is not reversed in high concentrations of extracellular Ca2+ ([Ca2+]o) as would be predicted by a direct interaction with Ca2+ channels. Cyproheptadine (0.5-1 microM), a serotonin antagonist that is structurally similar to amitriptyline, causes similar effects as reported above for the TCAs; however, ketanserine (10 microM), also a serotonin antagonist but without the tricyclic nucleus, is less effective in this regard. Carbamazepine, an anticonvulsant with a tricyclic nucleus, produces similar effects as the above three compounds only in higher, yet therapeutic, concentrations (50 microM). Neither 5-hydroxytryptamine nor norepinephrine (100 microM, each) had effects on the EAA-induced elevation of [Ca2+]i. This is the first report to show an interaction of tricyclic antidepressants with the function of glutamate receptors in concentrations which are consistent with therapeutic dosages.


Analysis of the relaxant action of SDZ PCO 400 in airway smooth muscle from the ox and guinea-pig. SDZ PCO 400 (30 nM-100 microM) suppressed the spontaneous tone of guinea-pig isolated trachealis. Glibenclamide (1-10 microM), phentolamine (100 microM), guanethidine (50 microM) and bretylium (50 microM) each antagonized SDZ PCO 400 without antagonizing isoprenaline or theophylline. Charybdotoxin (100 nM) failed to antagonize SDZ PCO 400 but antagonized theophylline. The relaxant action of SDZ PCO 400 was ablated when spasm was induced by a K(+)-rich (120 mM) medium. In bovine and guinea-pig trachea, SDZ PCO 400 (10 microM) suppressed spasm evoked by lower (less than 40 mM) but not higher (greater than 40 mM) concentrations of KCl. In guinea-pig trachea the relaxant action of SDZ PCO 400 was associated with suppression of electrical slow waves and with marked cellular hyperpolarisation. SDZ PCO 400 (0.5 and 10 microM) promoted the efflux of 86Rb+ from bovine trachealis, an effect inhibited by glibenclamide (1 microM). It is concluded that the tracheal relaxant action of SDZ PCO 400 is associated with the opening of a plasmalemmal K(+)-channel analogous to the ATP-sensitive K(+)-channel observed in insulin-secreting cells.


Effects of clorgyline on the release of noradrenaline from rat vas deferens. Release of endogenous noradrenaline (NA) and functional activity of presynaptic alpha 2-adrenoceptors were measured in isolated rat vas deferens following acute (2 mg/kg) or chronic (21 * 1 mg/kg daily, i.p.) treatment with clorgyline. Noradrenaline tissue content was higher in rats treated chronically with clorgyline than in those treated acutely. In vitro experiments done in the presence of 1 microM desipramine and 0.5 microM yohimbine showed that NA release, elicited by electrical field stimulation, was higher in chronic clorgyline rats than in controls, while no significant difference was found between acute clorgyline and control rats. Yohimbine enhanced evoked release of NA in all treatment groups, provided that desipramine was present in the Krebs solution, and the enhancement was non significantly higher in chronic clorgyline than in acute clorgyline and control rats. Efflux of 3,4-dihydroxyphenylglycol was lower in chronic clorgyline rats than in other groups. No difference was found between the treatment groups in a 1 min [3H]NA uptake into the tissue, nor in the ability of desipramine (1 microM) to block the uptake. The results indicate that following chronic treatment with clorgyline, evoked release of NA increased, and there was no reduction in the ability of the presynaptic alpha 2-adrenoceptor inhibitory mechanism to reduce nerve stimulation induced release of NA.


Characterization of the novel 5-HT3 antagonists MDL 73147EF (dolasetron mesilate) and MDL 74156 in NG108-15 neuroblastoma x glioma cells. In radioligand binding experiments, MDL 73147EF and MDL 74156 inhibited the binding of [3H]GR65630 to 5-hydroxy-tryptamine3 (5-HT3) binding sites on membranes prepared from NG108-15 neuroblastoma x glioma cells. The calculated dissociation constants (KI) were 20.03 +/- 6.58 and 0.44 +/- 0.18 nM, respectively (means +/- S.E.M., n = 6 and 9, respectively). Application of 5-HT (10-50 microM) to voltage-clamped NG108-15 cells elicited a rapidly desensitizing inward membrane current, characteristic for the activation of 5-HT3 receptors. The 5-HT-induced membrane current was suppressed in a reversible, concentration-dependent manner by MDL 73147EF and MDL 74156EF. The concentrations required to block half the 5-HT response (IC50) were 3.8 and 0.1 nM, respectively. It is concluded that both compounds are potent and reversible antagonists at 5-HT3 receptors in this neuroblastoma cell line.


Eicosanoids modulate CR1- and Fc-dependent bacterial phagocytosis. It is known that macrophages produce large amounts of eicosanoids during phagocytosis and that pharmacological concentrations of prostaglandin E2 (PGE2) inhibit phagocytosis in several models. However, the physiological effect on phagocytosis of endogenous prostaglandins, produced during CR1- or FcR-mediated bacterial phagocytosis, remains unclear. In this study, we show that indomethacin inhibits the CR1- but not the FcR-dependent phagocytosis of bacteria by rat peritoneal cells in the same range of concentrations that inhibit the synthesis of PGE2, PGI2 and thromboxane A2. An exogenous supply of PGE2 and PGE1 (10(-10) to 10(-8) M) restored the CR1-mediated phagocytosis; higher concentrations were inhibitory. Our data indicate that PGE2 and/or PGI2, produced by rat peritoneal cells, are involved in CR1-dependent bacterial phagocytosis.


Conversion of DNA adducts of antitumour cis-diamminedichloroplatinum(II). Immunochemical analysis. Polyclonal antibodies that bind selectively to DNA modified by antitumour cisplatin and its analogues were isolated. The reactivity of the antibodies with the epitope was enhanced by thermal denaturation of DNA that had been modified by cisplatin before its denaturation. On the other hand, denaturation of DNA before its modification resulted in considerably less reaction of the antibodies. The conversion of monofunctional cisplatin-DNA adducts to bifunctional lesions increased the capability of the modified DNA to competitively inhibit the antibodies. The double-helical oligonucleotides containing a unique bifunctional adduct formed by cisplatin at the d(GG) site cross-reacted with the antibodies in contrast to the oligonucleotide containing a single monofunctional adduct formed at the d(G) site. In addition, poly(dG-dC) . poly(dG-dC) modified by cisplatin did not react with the antibodies. It was concluded that the antibodies recognized monodentate lesions, intrastrand cross-links between two purine nucleosides separated by one or more nucleosides and interstrand cross-links negligibly. The antibodies apparently recognized a chemical nature of the bifunctional adduct formed between two adjacent purines and not an unusual conformational feature of DNA resulting from the formation of this adduct. The antibodies were used to analyse the adducts formed by cisplatin on DNA of cultured cells exposed to this drug. During the subsequent incubation of the already exposed cells in the drug-free medium, a part of the bifunctional adducts of cisplatin was completely removed from DNA or transformed to the adducts not recognized by the antibodies.


Characterization of ouabain high-affinity binding to rat cerebral cortex. Modulation by melatonin. High-affinity [3H]ouabain binding to membrane preparations of rat cerebral cortex was examined using a rapid filtration procedure. At 37 degrees C, binding reached equilibrium in about 60 min. Scatchard analyses of the data at equilibrium revealed a single population of binding sites with a dissociation constant of KD = 3.1 +/- 0.36 nM and a binding site concentration of Bmax = 246.4 +/- 18.4 fmol/mg protein. Kinetic analyses of the association and dissociation curves indicated a kinetic KD = 4.6 nM, which is in good agreement with the value obtained at equilibrium. When various digitalis compounds were tested for their ability to inhibit [3H]ouabain binding, the following Ki values (nM) were obtained: ouabain (3.9); digoxin (18); acetyl-digitoxin (66); k-strophanthin (95); digitoxin (236). When melatonin was added to the incubation medium, the ability of ouabain to inhibit [3H]ouabain binding increased in a dose-related manner to yield the following Ki values (nM): melatonin 10 nM (2); melatonin 20 nM (1.2); melatonin 40 nM (0.8). These data suggest the existence in the rat cerebral cortex of high-affinity ouabain binding sites which may be a locus for the molecular action of melatonin.


Effect of amphotericin B on intracellular calcium levels in cultured glomerular mesangial cells. Amphotericin B induces a reduction in the glomerular filtration rate due, in part, to a decrease in the glomerular capillary ultrafiltration coefficient (Kf), possibly resulting from mesangial cell contraction. The present study has examined the effect of amphotericin B on intracellular free calcium concentrations ([Ca2+]i) in cultured glomerular mesangial cells, using the fluorescent probe, fura-2. Under control conditions, amphotericin B caused a concentration-dependent increase in [Ca2+]i with a 231 +/- 52 nM increase at a concentration of 10(-6) M. This increase was almost completely inhibited when Ca2+ ions were omitted from the cell medium (an increase of 34 +/- 7 nM, P less than 0.0001). Replacement of extracellular Na+ ions with N-methylglucamine caused a marked inhibition of the amphotericin B-induced rise in [Ca2+]i (28 +/- 5 nM, P less than 0.0001). Diltiazem (20 microM) also suppressed the rise in [Ca2+]i seen with amphotericin B (36 +/- 6 nM, P less than 0.0001). In contrast, theophylline (20 microM) enhanced the response to amphotericin B (351 +/- 51 nM rise, P less than 0.001). The results suggest that amphotericin B-induced mesangial cell contraction is secondary to Ca2+ entry from the extracellular space through voltage-dependent calcium channels. The dependence of the response on extracellular Na+ suggests that Na+ entry induces depolarization of the membrane and opening of voltage-dependent calcium channels.


ADP-ribosylation of rho proteins is inhibited by melittin, mast cell degranulating peptide and compound 48/80. The amphiphilic agents melittin, mast cell degranulating peptide and compound 48/80 inhibit the ADP-ribosylation of the small GTP-binding proteins rho by Clostridium botulinum exoenzyme C3. Half-maximal and maximal inhibition (greater than 90%) of ADP-ribosylation occurred at about 8 and 25 micrograms/ml for compound 48/80, at 10 and 45 microM for mast cell degranulating peptide and at 15 and 50 microM for melittin, respectively. In addition, these compounds increase the steady state GTP hydrolysis and the association and dissociation rate of GTP-binding of rho proteins through an increase of GDP/GTP exchange. The data suggest that the amphiphilic agents tested interact with small GTP-binding proteins of the rho protein family.


Serotonin inhibits adenylate cyclase in human platelet membranes. Serotonin (5-HT), the non-specific 5-HT1 receptor agonist methysergide, and the 5-HT1B receptor agonist CGS 12066 dose-dependently inhibited forskolin-stimulated adenylate cyclase activity in membranes derived from human platelets. Buspirone and ipsapirone showed partial agonist activity. The effect of 5-HT to inhibit forskolin-stimulated activity was partially antagonised by spiperone, yohimbine and pindolol, with each of these compounds showing some partial agonist activity. It is concluded that human platelet membranes possess receptors for 5-HT coupled negatively to adenylate cyclase. The relationship of these receptors to other binding sites which have been shown for 5-HT on platelet membranes remains to be finally elucidated.


Stereoisomerism and muscarinic receptor agonists: synthesis and effects of the stereoisomers of 3-[5-(3-amino-1,2,4-oxadiazol)yl]-1- azabicyclo[2.2.1]heptane. The preparation and the biological activities of the four stereoisomers of 3-[5-(3-amino-1,2,4-oxadiazol)yl]-1-azabicyclo[2.2.1]heptane are described. The most potent stereoisomer, 3a, has the 3R,4R configuration, and in vitro activities in (pD2(% efficacy): ileum 8.8 (87%), hippocampus 9.8 (116%) and ganglion 10.2 (36%)). 3b (3S,4S) was weaker (ileum 8.1 (121%), hippocampus 8.5 (107%), ganglion 9.0 (63%)). The other two stereoisomers, 4a (3S,4R; ileum 7.1 (108%), hippocampus 8.2 (116%), ganglion 7.3 (31%)) and 4b (3R,4S; ileum 7.0 (100%), hippocampus 7.0 (120%), ganglion 7.2 (67%)) are of comparable activity, with an analogous profile to that of the more potent stereoisomers. Thus, compounds 3a and 4a, possessing the 4R stereochemistry, showed selectivity for the hippocampus over the ileum. Compound 3a was, however, more potent in the ganglion than in the hippocampus. All four stereoisomers were full agonists in the hippocampus, indicating M1 activity; however, they were partial agonists in the depolarisation of the rat superior cervical ganglion, another M1-mediated response. This may be due to M2-mediated hyperpolarization. With 3a (0.01 mg/kg i.p.), expression of c-fos mRNA was observed in the hypothalamus and in brain areas involved in sensory processing; these effects were totally blocked by pretreatment with 2 mg/kg scopolamine. In particular, activation of the superior colliculus is consistent with potent M2 activity.


Molecular modelling of asperlicin derived cholecystokinin A receptor antagonists. The C3-substituted benzodiazepines derived from asperlicin, e.g. devazepide (L-364,718, MK-329), constitute the most potent class of cholecystokinin A-type (CCKA) receptor antagonists. In order to gain insight into the prerequisites for binding, we examined the conformational properties of both potent and weak representatives of this class with computer assisted molecular modelling (CAMM) techniques. The CAMM results indicate that the binding site for the C3-substituents is a planar slot on the CCKA receptor surface and, in addition, allow the proposal of a model which describes the relative binding mode of the less potent R isomers versus that of the S isomers. The latter model illustrates the unique spatial properties of the benzodiazepine moiety, which we suggest functions primarily as an invertible core which assures an optimal arrangement of attached substituents.


Discrimination between two types of P2 purinoceptors by suramin in rat hepatocytes. Suramin, currently reported as a P2 purinoceptor antagonist, competitively inhibited P2 purinoceptor agonist-induced phospholipase C (PLC) stimulation, but not P2 purinoceptor agonist-induced adenylate cyclase (AC) inhibition in rat hepatocytes. Suramin did not inhibit vasopressin-induced PLC activity. We conclude that there are two types of P2 purinoceptors; one is suramin-sensitive and coupled to PLC in a stimulatory manner, and the other is suramin-insensitive and coupled to AC in an inhibitory manner.


Adenosine A1 receptor gene structure and regulation in normotensive and spontaneously hypertensive rats. In spontaneously hypertensive rats (SHR), both the response to adenosine and the affinity of adenosine A1 receptors (A1R) are altered. We compared the coding sequences and the mRNA expression levels of A1R in SHR and normotensive Wistar rats (NWTR). Neither the nucleotide sequence nor the mRNA level of A1R are altered in SHR, so that gene mutations or an altered gene regulation of A1R cannot account for alterations in A1R function in SHR.


Inhibition of human liver steroid sulfotransferase activities by drugs: a novel mechanism of drug toxicity? The inhibition of steroid and phenol sulfotransferase activities in human liver cytosol by a wide range of commonly used drugs was studied. Dehydroepiandrosterone (DHEA) and estrone sulfotransferase activities were strongly inhibited by a number of compounds, with IC50 values ranging between 440 pM and 147 microM. For DHEA sulfotransferase, clomiphene, testosterone, danazol and spironolactone were the best inhibitors, with IC50 values less than 5 microM, whereas for estrone sulfotransferase cyclizine, ibuprofen, chlorpheniramine and dimenhydrinate resulted in the strongest inhibition, again with IC50 values of less than 5 microM. The xenobiotic substrate 1-naphthol was refractory to substantial inhibition, with the exception of clomiphene. The majority of the drugs which inhibited steroid ST activities strongly were either synthetic steroids, antisteroidals or were tertiary amine drugs such as tricyclic antidepressants and antihistamines, many of which exhibit adverse side effects manifesting particularly as sexual dysfunction and disruption of hormone action in clinical use. The importance of these findings for our understanding of the molecular basis of adverse drug reactions is discussed.


Dietary iron lowers the intestinal uptake of cadmium-metallothionein in rats. It has been shown that addition of extra calcium/phosphorus (Ca/P), zinc (Zn) and iron (Fe2+) to the diet results in a significant protection against cadmium (Cd) accumulation and toxicity in rats fed inorganic Cd salt. However, it is not clear whether the presence of these mineral supplements in the diet also protects against the Cd uptake from cadmium-metallothionein. The present study examines the influence of Ca/P, Zn and Fe2+ on the Cd disposition in rats fed diets containing either 1.5 and 8 mg Cd/kg diet as cadmium-metallothionein (CdMt) or as cadmium chloride (CdCl2) for 4 weeks. The feeding of Cd resulted in a dose-dependent increase of Cd in intestine, liver and kidneys. The total Cd uptake in liver and kidneys after exposure to CdMt was lower than after exposure to CdCl2. At the low dietary Cd level and after addition of the mineral supplement, the kidney/liver concentration ratio increased. However, this ratio was always higher with CdMt than with CdCl2, suggesting a selective renal disposition of dietary CdMt. The uptake of Cd from CdCl2 as well as from CdMt was significantly decreased by the presence of a combined mineral supplement of Ca/P, Zn and Fe2+. The protection which could be achieved was 72 and 75% for CdMt and 85 and 92% for CdCl2 after doses of 1.5 mg/kg and 8 mg/kg respectively. In a following experiment it was shown that the protective effect of the mineral mixture against CdMt was mainly due to the presence of Fe2+. It seems clear that Cd speciation and the mineral status of the diet have a considerable impact on the extent of Cd uptake in rats.


Antihypercholesterolemic effect of dipyridamole in chickens. The administration of a 4 mg/kg dose of dipyridamole daily in chickens fed a diet supplemented with 2% cholesterol reversed the hypercholesterolemic effects of the diet. In particular, it reduced the plasma cholesterol concentration in approximately 18%; the levels of very-low-density lipoproteins and intermediate-density lipoproteins and the liver cholesterol content. Although the mechanism was not fully elucidated, the increased excretion of cholesterol seemed to be responsible for the lipid lowering effect. When dipyridamole was administered in chickens fed the same diet without cholesterol no significant changes were observed. Inasmuch as the chicken lipoprotein metabolism differs in several aspects to human, the extrapolation of the hypocholesterolemic effect of dipyridamole to man must be made with care.


Influence of age on epithelium-dependent responsiveness of guinea-pig and rat tracheal smooth muscle to spasmogens. The current study describes the influence of age and the presence of the epithelium on guinea-pig and rat tracheal airway smooth muscle sensitivity to the spasmogens histamine, acetylcholine, carbachol and potassium. In guinea-pig trachea from animals aged 2-52 weeks the potency of each of these spasmogens decreased with increased animal age. In contrast, no age-dependent changes in the potency of acetylcholine, carbachol or potassium were seen in rat trachea. Removal of the tracheal epithelium was associated with significant increases in the potencies of histamine and acetylcholine in guinea-pig trachea and of acetylcholine in rat trachea, but not of carbachol or potassium in either species. For histamine in guinea-pig trachea, the largest potency increase (4.6-fold) occurred in tissue from 6-week-old animals, with the smallest increases in tissue from the youngest (2 weeks) and the oldest (52 weeks) animals. Thus, although the sensitivity of airway smooth muscle to this spasmogen fell between 2 and 12 weeks of age, the effect of epithelial removal on sensitivity to histamine was apparently increased during this period. Further studies are required to assess the reasons for increased histamine and acetylcholine potency in airway smooth muscle after epithelial ablation.


DNA adducts in carp exposed to artificial diesel-2 oil slicks. In attempts to mimic field exposure, oil slicks prepared from diesel-2 oil/water emulsions were poured onto the surface of water in tanks prepared fresh every day and liver DNA adducts were analyzed by 32P-postlabeling in carp free-swimming in these tanks. 'Clusters' of lipophilic DNA adducts were detected, with five major and numerous minor adducts. Essentially a similar adduct pattern was found in the liver DNA of carp exposed to crude oil-polluted water. Diesel-2 adduct induction was observed slowly with a steady increase to greater than 3000 amol/microgram DNA at day 12. After this time fish were transferred to clean water. Adduct levels continued to increase through day 17 (approximately 10,000 amol/microgram DNA) despite the cessation of exposure, but a 30% and 80% decline was evident at day 22 and day 27, respectively. All major adducts were distinct from the known benzo[a]pyrene diolepoxide-dG. These results indicate that diesel-2 oil can cause extensive DNA damage in carp in vivo and the damage accumulates proportionately with time of exposure.


In vitro metabolism of isaxonine phosphate: formation of two metabolites, 5-hydroxyisaxonine and 2-aminopyrimidine, and covalent binding to microsomal proteins. Isaxonine phosphate or Nerfactor (2-isopropylaminopyrimidine) has been implicated in several cases of hepatitis which is reversible after withdrawal of the drug. In order to understand the cause of such hepatitis, the metabolic activation of isaxonine phosphate with different liver microsomes was investigated. The major metabolites were 5-hydroxyisopropylaminopyrimidine and 2-aminopyrimidine. Covalent binding to microsomal proteins was also detected. In vitro metabolic activation required intact microsomes, NADPH and O2 as cofactors and was cytochrome P-450 dependent. A sensitive fluorimetric assay of 5-hydroxyisaxonine was developed. The metabolism of isaxonine phosphate was compared in liver microsomes from rat, rabbit, dog, monkey and man and found to be qualitatively similar. Treatment of rats with phenobarbital increased the formation of 5-hydroxyisaxonine, while treatment with 3-methylcholanthrene increased the formation of 2-aminopyrimidine but decreased that of 5-hydroxyisaxonine. Inhibition and reconstitution experiments demonstrated that 5-hydroxylation of isaxonine was catalyzed by a cytochrome P-450. Metabolic oxidation of isaxonine phosphate using 5-[3H]isaxonine phosphate led to a total loss of tritium in 5-hydroxyisaxonine and partial loss of tritium in 2-aminopyrimidine and covalent binding to proteins.


Routes for renal transport of methylmercury in mice. To clarify the routes for renal methylmercury uptake, the effects of ureter ligation and pretreatment of probenecid, an organic anion transport inhibitor, or acivicin, a gamma-glutamyltranspeptidase (gamma-GTP) inhibitor, on renal methylmercury content were investigated in mice. For 120 min after CH3HgCl (5 mumol/kg, i.v.) injection, renal methylmercury content in bilateral ureter-ligated mice was approximately 50% lower than that of sham-operate mice. The glomerular filtration rate was reduced to about 15% of the control by ureter ligation. These results suggest an important role of glomerular filtration in the renal methylmercury uptake. Pretreatment with probenecid (0.5 or 1.0 mmol/kg, i.p.) reduced the renal methylmercury accumulation 30 min after CH3HgCl injection in a dose-dependent manner in both ureter-ligated and sham-operated mice. Urinary methylmercury excretion was not affected by probenecid pretreatment. Renal methylmercury content of ureter-ligated mice was not changed by pretreatment with acivicin (0.5 or 1.0 mmol/kg. i.p.), which was previously reported to decrease the renal methylmercury content in mice. Coadministration of GSH (10 mumol/kg, i.v.) with CH3HgCl increased the renal methylmercury uptake determined 5 min after injection in ureter-ligated mice. These results suggest that at least two transport systems play major roles in renal methylmercury uptake: one is a route from the glomeruli through the brush border membrane which is dependent on the action of gamma-GTP, and the other route is the one using an organic anion transport system through the basolateral membrane.


Immunohistochemical localization of epidermal growth factor and its receptor during odontogenesis in the rat. The expression of epidermal growth factor (EGF) and epidermal growth factor receptor (EGFr) in developing teeth has been immunohistochemically studied in rat embryos (E-16 to E-21). Both EGF and EGFr showed a similar pattern of distribution. A very weak immunostaining was observed in the dental germ cells during the bud, cap, and bell teeth stages, as well as in few ectomesenchymal cells. In developed, but not erupted teeth, a moderate immunoreactivity for EGF and EGFr was present in the odontoblasts, in the ameloblasts and in the internal epithelial cells, but it was stronger in the dentine. In addition, the presence of EGF/EGFr was also observed in the intercalated ducts of salivary glands, primarily the submaxillary gland, in the maxillary bone cells, and in the cells of the peripheral and central nervous system. These results suggest that EGF has little or no effect during the early periods of tooth differentiation, whereas it is probably involved in the production of dentine. Moreover, EGF/EGFr seem to participate in the maturation and differentiation of other embryonic tissues such as tissues of the nervous system and bone.


Muscle response to the oral-screen activator. An EMG study of the masseter, buccinator, and mentalis muscles. The myofunctional changes that occur during orthodontic treatment with functional appliances constructed with vestibular screen elements, such as buccal shields and lip pads, have not been sufficiently investigated. The aim of this study was to examine electromyographically the response of the masseter, buccinator and mentalis muscles to the oral-screen activator (OSAC), which is a conventional activator constructed with buccal shields and lip pads. The material consisted of 10 children, with Angle Class II, division 1 malocclusion and narrow dental arches (mean age 10.7 years), treated at the Department of Orthodontics, School of Dentistry, Malmö, Sweden. The group had a mean overjet of 8.5 mm, and a mean ANB angle of 6 degrees. Six out of the 10 cases had incompetent lips at relaxed lip position. The construction bite of the OSAC was taken with the anterior teeth in edge-to-edge position. The recording procedure included the following positions: (a) postural position of the mandible with lips relaxed and lips closed; (b) clenching; (c) swallowing saliva; and (d) spatula exercise. EMG recordings were made after 1, 2, and 3 months of appliance treatment. Differences between means were tested with a two-way analysis of variance (ANOVA). The results showed that during postural position with lips closed there was a statistically significant increase in mentalis EMG with the oral-screen activator in place. The buccinator and masseter muscles showed no EMG activity during rest with or without the appliance in the mouth. During clenching we found no statistically significant difference in masseter EMG with or without the appliance in place. During swallowing saliva a statistically significant decrease was recorded in mentalis EMG with the oral-screen activator in the mouth, while buccinator activities remained the same with and without the appliance. The masseter activity, on the other hand, increased significantly with the activator in place. During spatula exercise mentalis and buccinator were highly activated. The conclusions drawn from this study are that the oral screen activator (OSAC) increased masseter activity during swallowing, but it remained unchanged during clenching. Lip pads increase mentalis activity during lip closure, but reduce mentalis hyperactivity during swallowing. The buccinator activity was insignificant and buccal shields did not change that activity. Spatula exercise stimulates both mentalis and buccinator activity.


Perception of malocclusion in 11-year-old children: a comparison between personal and parental awareness. In orthodontic counselling an understanding of how individuals perceive their occlusal features is important to ensure effective communication and for provision of adequate advice on treatment need. The purpose of the present study was to assess personal and parental awareness of malocclusion in children, and to examine whether agreement existed between children and their parents on assessments of malocclusion traits. Of 104 randomly selected fourth-grade children 99 presented themselves at a public dental clinic. Ninety-three accompanying parents attended. Awareness was assessed by comparing the opinion of parents and children on the presence or absence of anterior malocclusion with direct measurements on dental study casts. The subjects' abilities to identify a polaroid of the child's dentition in a panel of 17 alternative photographs were also used as a measure of awareness. The findings revealed a moderate level of awareness among both the children and their parents. About half of the children and the parents identified the child's photograph on the first attempt. About three-quarters of the traits recorded as marked/severe malocclusion and about half of the mild/moderate traits were recognized. A significant association existed between the number of correct reports on traits given by the children and the parents. However, agreement across professional, child, and parental assessments varied for the different traits. The results indicated that the individual's comprehension of professional terms may be unclear and that professionally defined cut-off points often do not coincide with norms existing within the actual family unit.