Behavioral effects of propentofylline and N6-(L-2-phenylisopropyl)-adenosine (PIA) were evaluated by operant behavior under a punishment situation, discrete shuttle avoidance response and ambulatory activity in mice. Propentofylline (3 mg/kg, s.c.) and PIA (0.01 mg/kg, s.c.) significantly decreased the punished response without producing a significant change in the non-punished response. Propentofylline and PIA reduced the increase in the punished response induced by caffeine (30 mg/kg) and diazepam (1 mg/kg). Propentofylline and PIA also reduced the increase of ambulation induced by caffeine. Furthermore, the single administration of propentofylline and PIA decreased the response rate and/or % avoidance in the discrete shuttle avoidance situation. However, the effective doses of propentofylline and PIA to reduce the ambulation-increasing effect of caffeine and to produce a change in the avoidance behavior were much higher than those effective for eliciting a significant change in the punished response. The present results suggest that there is an intimate interaction between central adenosine and benzodiazepine systems with regards to the change of punished response.
Vasomotor effects of histamine were examined in isolated coronary arteries from pigs and cattle. Histamine produced a concentration-dependent contraction in these arteries. These contractile responses were dose-dependently inhibited by diphenhydramine. The slopes of the Schild plots, however, were significantly lower than unity in both species. Cimetidine potentiated the histamine-induced contractions at relatively high doses of histamine (larger than 10-5 M) in pig coronary arteries, but did not show a significant effect in cattle arteries. After the removal of endothelium, the Schild plot of diphenhydramine against histamine gave a straight line with a pA2 value of 7.80 and slope of 1.00 in pigs, confirming the competitive nature of the antagonism. In cattle, the slope was significantly lower than unity; however, in the presence of cimetidine, it was not significantly different from unity. Dimaprit did not contract the cattle coronary arteries with endothelium, but contracted them after the removal of endothelium. These results suggest that histamine-induced vasoconstriction in pig and cattle coronary arteries is mainly dependent on the H1-receptors in the smooth muscle cells, and that H1- and H2-receptors in the endothelial cells of pigs and H2-receptors in the smooth muscle cells of cattle modify the histamine-induced vasoconstrictions.
The effects of flutropium on histamine (Hist)-induced increase in intra nasal pressure in non-sensitized guinea pigs and nasal mucosa capillary permeability in passively sensitized guinea pigs were investigated. Flutropium (0.3%), atropine (0.3%), diphenhydramine (0.01%) and cimetidine (0.1%) were directly inhaled into the nasal cavities by an ultrasonic nebulizer for 20 min, followed by inhalation of Hist (0.1%) for 10 min. Flutropium, atropine and diphenhydramine had an inhibitory action on the Hist-induced increase in intranasal pressure in guinea pigs. Cimetidine had no effect on this system. In passively sensitized guinea pigs (the challenge was performed 48 hr after sensitization), a 0.1-1.0 mg/kg injection of flutropium (i.v.) dosedependently inhibited the allergic nasal mucosa capillary permeability. Atropine (10 mg/kg, i.v.) had no inhibitory action on this system. These results suggest that inhalation into the nasal cavities and i.v. injection of flutropium are effective in experimental models of drug and allergy-induced rhinitis of the guinea pig.
From functional studies with propylbenzilylcholine mustard (PrBCM), we reported that there coexist PrBCM-sensitive and PrBCM-resistant muscarinic cholinoceptor mechanisms in guinea pig taenia caecum. We investigated the interrelationship between these two cholinoceptor mechanisms using an in vitro receptor binding assay with [3H]quinuclidinyl benzilate (QNB) and [3H]PrBCM. Pretreatment of the muscle strips with 300 nM PrBCM (in vivo alkylation) for 10-50 min resulted in progressive decreases of the number of the maximum [3H]QNB binding sites. However, a prolongation of the period of in vivo alkylation up to 90 min was accompanied with no further loss in the binding sites. Under these conditions, there is no significant change in the affinity of [3H]QNB for the binding sites. The concentration of carbachol required to displace 50% of the bound [3H]QNB was larger in membranes obtained from the tissues that had been alkylated in vivo with PrBCM for 50 min than that from control strips, but was not altered when the pretreatment with the drug was carried out after homogenization (in vitro alkylation). When GTP was added during in vitro alkylation, the affinity of carbachol was lower than that in control membranes, as observed when in vivo alkylation was carried out. In the presence of guanine nucleotide, PrBCM thus appears to recognize two distinct populations or states of muscarinic receptors.
The effect of in vitro exposure of sarcolemmal membrane (SL) vesicles to Gram-negative endotoxin lipopolysaccharides (LPS) was studied. LPS decreased the Na, K-ATPase activity of SL vesicles; this effect was inhibited by hydroxyl radical (·OH) scavengers such as dimethylthiourea and dimethyl sulfoxide, but not by superoxide dismutase, a scavenger of superoxide anion radicals or by the hydrogen peroxide scavenger catalase. ESR spin-trapping with 5, 5-dimethyl-l-pyrroline N-oxide verified the generation of ·OH from LPS itself under the conditions used; ·OH gener ated from LPS was not affected by deferoxamine, a powerful iron chelator. The Na, K-ATPase activity was reduced by an ·OH radical generating system consisting of dihydroxyfumarate and Fe3+-ADP. Furthermore, exposure of SL vesicles to LPS caused an increase in malondialdehyde formation. It can be concluded that LPS damages cardiac SL by an oxygen free radical mechanism by the generation of ·OH, due to inhibition of Na, K-ATPase activity and peroxidation of lipids, and that the effect of LPS is not dependent on the presence of contaminating iron.
The effects of betaxolol on isolated rat arteries and the modes of action were investigated. Betaxolol (10-5 - 10-3M) relaxed the 80 mM K+-induced contraction of aortic strips concentration-dependently. The 50% inhibitory concentration of betaxolol in the K+-induced contraction was 3 times higher than that of papaverine and about 3 times lower than that of bunitrolol. The relaxations by betaxolol were also demonstrated in renal, mesenteric and femoral arteries. Betaxolol (3 × 10-6M - 10-4M) produced rightward parallel shifts of the concentration-response curves for Ca 2+ in the K+-depolarized aortic strips. On the other hand, betaxolol produced downward shifts as well as rightward shifts of the concentration-response curves for norepinephrine, 5-HT and angiotensin II. In K+-depolarized aortic strips, the cytosolic Ca2+ concentration measured with a fluorescent indicator, fura-2, was decreased by betaxolol (10-4M) almost concomitantly with the loss of tention. An elevation of external Ca2+ from 2.5 mM to 10 mM restored both the cytosolic Ca2+ concentration and tention. The relaxations of arteries induced by betaxolol were not influenced by glybenclamide, methylene blue, indomethacin or removal of the endothelium. These results suggest that betaxolol possesses a direct vasodilating action, and the action may be due to the inhibition of Ca2+ influx across the cell membrane.
Experiments were performed on spinalized rats transected at C1. Intravenous administration of 2-phenylethylamine-HC1 (PEA-HC1) (0.3 and 1 mg/kg, i.v.) and methamphetamine-HC1 (MAP-HC1) (0.1 and 0.3 mg/kg, i.v.) increased the amplitude of the monosynaptic reflex (MSR). The increase of the MSR caused by PEA and MAP was antagonized by prazosin-HC1 and abolished by the pretreatment with reserpine (i.p.) and 6-hydroxydopamine (intracisternally, 14 days previously). A dopamine D1 antagonist, SK&F 83566-HBr (0.01 mg/kg, i.v. ), and a D2 antagonist, YM-09151-2 (0.3 mg/kg, i.v.), did not antagonize the increasing effects produced by PEA and MAP. An inhibitor of type-B monoamine oxidase, (—)deprenyl-HCl (1 mg/kg, i.v.), prolonged the effect of PEA but not that of MAP, suggesting that PEA alone, and not its metabolites, enhanced the MSR. These results suggest that PEA and MAP increase the amplitude of the MSR by releasing noradrenaline from the terminals of descending noradrenergic fibers, and that PEA, an endogenous trace amine, has a mechanism of action similar to that of MAP.
The spike after-hyperpolarization (AH), which is linked to the Ca-activated K conductance system, in the rat superior cervical ganglion was examined by means of intracellular recording. The AH was shortened during repetitive stimulation of the cell at 1 Hz and reached a steady state within 10 sec. These changes in the AH disappeared after a part of the AH was depressed by 5μM ryanodine. The steady state of AHs was dependent on the frequency of stimulation ranging from 0.005 Hz and 2 Hz; half depression was observed at 0.53 Hz. Half recovery time from the depression induced by conditioning stimuli at 2 Hz was approximately 14 sec. Caffeine (1 mM) or TEA (3 mM) enlarged the AH at low stimulation frequencies. Caffeine slightly shifted the frequency depressing the AH toward high frequencies, and TEA did not cause significant changes. These results suggest that the repetitive firing of cells decreases the intracellular Ca release by contributing to the generation of the AH, rather than by accelerating the loading of Ca to its storage sites.
To clarify the mechanisms underlying the vasoactive effects of endothelin-3 (ET-3), we examined the effects of indomethacin, endothelium-denudation, methylene blue and L-NG-monomethyl arginine (L-NMMA) on the perfusion pressures of isolated rat mesenteric arteries infused with ET-3. ET-3 at 10-15 10-8 M elicited significant vasodilations in a dose-related manner, in which 10-9 and 10-8 M ET-3 caused biphasic pressure changes involving a transient dilation and subsequent vasoconstriction. Five micromolar indomethacin did not affect the vasodilations and vasoconstrictions induced by ET-3. In endothelium-denuded arteries, 10-13-10-8 M ET-3 elicited significant vasoconstriction in a dose-related manner without any vasodilation. In the presence of 30μM methylene blue, the vasodilations induced by ET-3 disappeared. In the presence of 100μM L-NMMA, 10-15-10-8 M ET-3 elicited significant vasoconstrictions in a dose-related manner, and the vasodilation by ET-3 existed only at 10-8M ET-3. These data suggest that the vasodilating effects of low doses of ET-3 through the endothelium overcome the vasoconstricting effects, and that the vasodilating effects of ET-3 are associated with an endothelium-derived relaxing factor as an endothelium-derived nitric oxide.
Stimulation of discrete intracardiac sympathetic nerves to the SA (SAS stimulation) or AV nodal region (AVS stimulation) increased the heart rate or decreased AV conduction time and caused an AV junctional rhythm, respectively, in anesthetized dogs treated with atropine. Topical application of tetrodotoxin (TTX) at the SAS or AVS stimulation locus totally inhibited the response to each stimulation, whereas each TTX treatment slightly attenuated the chronotropic response to the right ansa stimulation by 23 ± 7.7% and the dromotropic response to the left ansa stimulation by 7 ± 7.5%. TTX abolished AVS stimulation-induced one. Before atropine, topical application of hexamethonium at the locus for stimulation of intracardiac parasympathetic nerves to the SA (SAP stimulation) or AV nodal region (AVP stimulation) abolished almost totally negative chronotropic responses to SAP and cervi cal vagus stimulation or negative dromotropic responses to AVP and cervical vagus sti mulation, respectively. These results demonstrate that activation of a very small population of intracardiac sympathetic nerves to target cells is enough to induce positive chronotropic and dromotropic responses in the heart in situ, and that SA and AV nodal pacemaker activity and AV conductivity are controlled multi-directionally by in tracardiac sympathetic nerves in contrast with parasympathetic ones.
The pharmacological profile of a new alpha-adrenoceptor antagonist, KT-611 (naftopidil), was studied in vitro. In the dog mesenteric and carotid arteries and in the rabbit, guinea pig and rat thoracic aortae, KT-611 competitively inhibited α1-adrenoceptor-mediated contractions induced by noradrenaline with pA2 values ranging from 6.73 to 8.15. KT-611 also inhibited the postjunctional α2-adrenoceptormediated contractions in the dog saphenous vein (pA2 = 6.77) or dog basilar artery. However, the responses mediated through prejunctional α2-adrenoceptors (rat vas deferens), β-adrenoceptors (rat atria), muscarinic receptors (guinea pig ileum) and 5-HT2 receptors (dog mesenteric artery) were little affected by KT-611. KT-611 also inhibited the sympathetic adrenergic contraction evoked by electrical transmural stimulation in the dog mesenteric artery, and the inhibition was not relieved upon repetitive washing for 1 hour with the drug-free solution. 3H-prazosin and 3H-clonidine binding to the rat cortex membranes was inhibited by KT-611 with pKi values of 7.69 and 5.75, respectively. These results suggest that KT-611 is an α1-adrenoceptor antagonist with a weak antagonistic activity to postjunctional α2-adrenoceptors.
Binding of [3H]guanosine triphosphate (GTP) with a high affinity was found to be present in the coated vesicle fraction prepared from bovine cerebral cortex. The binding was saturable and displaced by 1μM of GTP, guanosine diphosphate and guanosine 5'-(3-O-thio)triphosphate. Incubation of the vesicles with islet-activating protein and [32P]NAD resulted in ADP-ribosylation of a 39, 000—41, 000-dalton polypeptide. Antibodies to the α-subunit of stimulatory guanine nucleotide regulatory proteins (G-proteins) immunoblotted 52, 000 and 45, 000-dalton polypeptides. The results indicate that stimulatory and inhibitory G-proteins are contained in a fraction of the bovine brain coated vesicles.
The effects of L-tryptophan on the development of tolerance to the antitussive effects of dihydrocodeine were examined in rats. Chronic co-administration of L-tryptophan with dihydrocodeine prevented the development of tolerance to the antitussive effects of dihydrocodeine. Furthermore, the antitussive effects of dihydrocodeine in dihydrocodeine-tolerant rats were fully restored by acute co-administration of L-tryptophan, as evidenced by a decrease of about 50% in the ED50 value of dihydrocodeine. Thus, it is concluded that L-tryptophan may antagonize the development of tolerance to the antitussive effects of dihydrocodeine.