Different effects of neuropeptide Y on electrically induced contractions in the longitudinal and circular smooth muscle layers of the female rabbit urethra

The effects of neuropeptide Y (NPY) on preparations of isolated longitudinal and circular smooth muscle from rabbit urethra were studied. In both types of muscle, electrically induced contractions and relaxations could be abolished by tetrodotoxin, (TTX). In the longitudinal muscle preparations the contraction was slightly reduced by prazosin, but markedly reduced by scopolamine and NPY. The NPY effect was not influenced by pretreatment with rauwolscine. Pretreatment with NPY had no effect on contractions induced by noradrenaline (NA) or carbachol and the peptide did not relax preparations contracted by these agents. In circular muscle an initial, fast response, not sensitive to prazosin or scopolamine was occasionally observed following electrical stimulation. A slow contraction component was regularly seen; this response was abolished by prazosin. Neuropeptide Y did not influence any of these responses. The preparations were concentration-dependently contracted by NA, whereas carbachol had no effect. Pretreatment with NPY did not affect contractions induced by NA, nor did the peptide relax NA-contracted preparations. In neither longitudinal nor circular muscle strips did NPY affect the electrically induced TTX sensitive relaxation of NA-contracted preparations. The results suggest that in the rabbit urethra NPY reduces contractions in the longitudinal muscle layer by selectively inhibiting the release of acetylcholine from cholinergic nerves. Neuropeptide Y did not appear to have any significant postjunctional effects nor to interfere with the release, or effects of NA or other transmitter agents. The physiological importance of the urethral effects of NPY remains to be established.     

Effects of some peptides on isolated human penile erectile tissue and cavernous artery

Contractant and relaxant effects of four peptides known to occur in nerves innervating human penile vessels and erectile tissue, namely substance P (SP), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY) and somatostatin, were studied in isolated preparations from the corpus cavernosum (CC), corpus spongiosum (CS) and cavernous artery (Acc). In addition, the actions of another peptide, arginine vasopressin (AVP), were investigated. In erectile tissue proper, SP induced concentration-dependent contractions. No effect of this peptide was observed in Acc segments. CC and CS preparations contracted by noradrenaline (NA) were relaxed by 30-40%; the effect in NA-contracted Acc preparations was inconsistent. AVP had a potent contractant effect in preparations from all the tissues studied, the effect being most conspicuous in CS strips. VIP was without contractant actions in any of the preparations. NA-contracted preparations were relaxed by VIP, and electrically induced contractions inhibited. The inhibitory effect was particularly marked in electrically stimulated CC and CS preparations. NPY had no effects; somatostatin contracted Acc segments, and in high concentrations CC and CS strips. It is concluded that among the peptides studied only VIP has effects compatible with a role as a neurotransmitter in penile erection.     

Effects of nimodipine, Bay K 8644 and pinacidil on alpha 1- and alpha 2-adrenoceptor-mediated vasoconstriction in human hand veins

The effects of nimodipine, Bay K 8644 and pinacidil, three drugs interfering with transmembrane Ca2+ fluxes in different ways, were investigated in isolated human hand veins. Their ability to influence the concentration-response relationship for noradrenaline (NA) was assessed in the absence and presence of prazosin or rauwolscine. The contractile response to NA was almost abolished in Ca2+ -free medium. Nimodipine and pinacidil depressed the NA concentration-response curve both in the absence and presence of alpha-adrenoceptor blockers. The NA response was only partially inhibited by nimodipine, indicating that NA may activate nimodipine-insensitive influx pathways, presumably receptor-operated calcium channels. Pinacidil inhibited the contractile response to 124 mM K+ and reduced the NA-induced contraction in the presence of nimodipine, suggesting that pinacidil has actions other than the opening of potassium channels and subsequent membrane hyperpolarization. Bay K 8644 increased the NA potency fourfold in the presence of rauwolscine, whereas it had no effect on the NA response in the presence of prazosin and in the absence of alpha-adrenoceptor blockade. Such an action of Bay K 8644 can be reconciled with alpha 1-adrenoceptor activation causing membrane depolarization and opening of potential-operated calcium channels. It may be concluded that both alpha 1- and alpha 2-adrenoceptor-mediated contractions in human hand veins are highly dependent on Ca2+ influx, although the mechanisms utilized to bring about this influx partly differ between the two receptor subtypes.     

Contractile effects of endothelin-1 and localization of endothelin binding sites in rabbit lower urinary tract smooth muscle

In isolated rabbit bladder and urethral smooth muscle, endothelin-1 caused concentration-related, slowly developing contractions that were difficult to wash out. Relative to contractions induced by K+ (124 mM), contractions in bladder preparations reached a higher amplitude than in urethral preparations. There was a marked tachyphylaxis to the effects of the peptide. The endothelin-1-induced contractions were not significantly affected by phentolamine or indomethacin in the urethra, or by scopolamine or indomethacin in the bladder. Incubation for 30 min in a Ca2(+)-free solution abolished the endothelin-1-induced contractions. Nifedipine did not affect the actions of endothelin-1 in the urethra but had a marked inhibitory action on its effects in the bladder. In the presence of endothelin-1, Ca2(+)-induced contractions were significantly blocked by nifedipine in the bladder but not in the urethra. Urethral preparations at resting tension responded to electrical stimulation by tetrodotoxin-sensitive, frequency-dependent contractions sensitive to alpha-adrenoceptor blockade. Pretreatment with endothelin-1 (10(-9) M) produced a significant increase in the nerve-induced contractions but had no significant effect on contractions induced by exogenous noradrenaline. Endothelin-1 did not affect spontaneous or stimulation-induced efflux of 3H-labelled noradrenaline in urethral smooth muscle. Preparations contracted by endothelin-1 were frequency-dependently relaxed by electrical stimulation. The peptide had no significant effect on the responses induced by electrical stimulation in the bladder preparations. In both bladder and urethra, [125]endothelin-1 binding sites were found mainly in the outer longitudinal muscle layer, in vessels and in the submucosa. The highest density of binding sites appeared to be in vessels and the outer muscle layer in both types of muscle. The results suggest that in the rabbit both bladder and urethral smooth muscle contain binding sites for endothelin. The peptide has contractant effects dependent on extracellular calcium in both types of tissue, but voltage-operated calcium channels seem to involved in activation only of bladder smooth muscle. The functional importance of endothelin-1 in the rabbit lower urinary tract remains to be elucidated.     

Vasodilator responses to alpha-human-atrial natriuretic peptide in isolated omental and pulmonary arteries from rabbit and man

In isolated vessels from the pulmonary and mesenteric/omental arterial circulations of rabbit and man, the effects of synthetic α-human atrial natriuretic peptide (α-hANP) were investigated. The vessels had an outer diameter of 0.6–1.2 mm and were contracted by noradrenaline (NA) and prostaglandin (PG) F_2α, (mesenteric/omental arteries) or by 5–hydroxytryptamine (5–HT) and PGF_2α (pulmonary arteries). It was found that in rabbits, mesenteric vessels contracted by NA and PGF_2α were not significantly relaxed by α-hANP. Rabbit pulmonary vessels contracted by PGF_2α or 5–HT were concentration-dependently relaxed by α-hANP (maximum 70%). Human omental vessels contracted by NA and PGF_2α showed a moderate (30%) relaxation after addition of the peptide. Human pulmonary arteries contracted by 5–HT were relaxed by α-hANP almost to baseline, while PGF_2α-contracted vessels showed a maximum relaxation of 65%. It is concluced that α-hANP has a relatively selective effect on pulmonary arterial vessels from rabbit as well as man. The results suggest that the peptide may be involved in the regulation of pulmonary vascular tone.     

Contractile effects of various vasoactive agents in small rat portal veins and hepatic arteries and the influence of sympathetic denervation on the noradrenaline response

Contractile responses were studied in isolated tubal segments of branches of the rat portal vein (luminal diameter approximately 300 microns) and hepatic artery (luminal diameter approximately 200 microns). Portal veins were approximately three times more sensitive to noradrenaline (NA) than hepatic arteries. 5-hydroxytryptamine contracted hepatic arteries concentration-dependently, whereas it produced only weak and inconsistent contractions in portal veins. Vasopressin effectively contracted hepatic arteries, whereas it had no effect on portal veins. Both vessel types responded to prostaglandin F2 alpha with contractions, although the drug potency was relatively low (EC50 greater than 10(-5) mol l-1). Histamine and carbachol failed to induce (hepatic arteries) or caused only weak (portal veins) contractions. Microsurgical hepatic hilar denervation reduced the catecholamine content of the parenchyma to less than or equal to 25% of controls. In both portal veins and hepatic arteries, the denervation procedure increased the NA sensitivity by factors of 3.1 and 2.0, respectively. In non-denervated livers, cocaine produced a similar increase of the NA sensitivity, whereas the drug had no significant effect in vessels from denervated animals. Thus, there was a marked difference between rat portal veins and hepatic arteries in their responsiveness to several contractile agents. Furthermore, the results of the present study indicate that the adrenergic nerves in both vessel types can be adequately removed by the microsurgical denervation procedure used.     

Characterization of the contractile effect of neuropeptide Y in feline cerebral arteries

The action of neuropeptide Y (NPY), which coexists with noradrenaline (NA) in perivascular sympathetic nerves, has been examined on feline cerebrovascular smooth muscle using a sensitive in vitro system. The direct cerebrovascular responses of peptides with structural similarities with NPY, peptide YY (PYY), avian (APP), and bovine (BPP) and human (HPP) pancreatic polypeptides, have been compared with that of NPY on isolated feline cerebral arteries. The relative potency for contractions induced by the peptides is: NPY, PYY greater than APP greater than BPP, HPP. The alpha-adrenoceptor antagonist rauwolscine, which blocked the response to noradrenaline (NA), had no effect on NPY-induced contractions. Neuropeptide Y significantly potentiated contractions induced by 10(-6) M NA, but not by 10(-5) M. Withdrawal of Ca2+ from the extracellular medium for 30 min reduced the contractile response to NPY in cerebral vessels by about 80%. Subsequent readdition of Ca2+ caused reproducible contractions which were inhibited by the calcium entry blocker nimodipine. Nimodipine also relaxed isolated middle cerebral artery segments contracted by NPY and NA in a concentration-dependent manner. The data suggest that NPY mediates contraction of cerebrovascular smooth muscle via a mechanism that is dependent on the concentration of extracellular calcium.     

Analysis of the effects of graded levels of hypoxia on noradrenaline-evoked contraction in the rat iliac artery in vitro

In rings of rat iliac artery, contractions were evoked by noradrenaline (NA), the selective alpha(1) adrenoceptor agonist phenylephrine (PE), and K(+), which causes depolarisation-induced contraction. There was no evidence of alpha(2) adrenoceptor-evoked contraction. Hypoxia, induced by reducing P(O(2)) in the bath from 100 mmHg to 70, 55 or 40 mmHg, had similar effects on rings with (E+) and without (E-) endothelium. In E- rings, the NA concentration-response curve was biphasic, whereas that for PE was monophasic. Hypoxia reduced maximum contractions in response to NA and PE (NA(max) and PE(max), respectively) without affecting the concentrations that evoked 50 % of maximum contraction (EC(50)). At P(O(2)) of 70 mmHg, NA(max) of the high affinity alpha(1) receptor for NA (NA(maxh)) and PE(max) were reduced by approximately 15 %, but at P(O(2)) of 55 and 40 mmHg, NA(maxh) was severely attenuated while PE(max) fell by 45 and 75 %, respectively. Similarly, the Ca(2+) channel blocker nicardipine depressed NA(maxh) and PE(max), but P(O(2)) of 55 mmHg further reduced NA(max) and PE(max). Hypoxia also reduced contractions evoked by NA, PE or K(+) at the concentrations required to produce 80 % of the maximum contraction (EC(80)), receptor-mediated contractions being more affected. Ca(2+)-free conditions reduced the contractions evoked by NA and PE, at the EC(80), to approximately 10 % of control. The K(+) channel inhibitors glibenclamide and tetraethylammonium did not prevent hypoxia-induced depression of PE-evoked contraction. Thus, contractions evoked in iliac artery by the high affinity subtype of alpha(1) adrenoceptor for NA, which may respond to circulating levels of NA, and by the single alpha(1) adrenoceptor subtype for PE, are especially vulnerable to P(O(2)) levels less-than-or-equal 55 mmHg. We propose that this reflects hypoxia-induced inhibition of Ca(2+) influx through L-type and receptor-operated Ca(2+) channels; K(+) channel opening makes little contribution.     

Role of Ca2+ in genesis of lower esophageal sphincter tone and other active contractions.

The effects of absent or low Ca2+ (0.5 mM), verapamil, nifedipine, Na nitroprusside, theophylline, La2+, and ethanol on basal active tension (tone), "off" contractions, and carbachol contractions were studied in opossum lower esophageal sphincter strips. Incubation in Ca2+-free Ringer (0.1 mM EGTA) abolished tone and contractions. Low Ca2+, verapamil, nifedipine, and theophylline depressed tone more rapidly than "off" contractions. Only verapamil and nifedipine depressed carbachol contractions. Na nitroprusside rapidly depressed tone but left contractions unchanged. La3+ at 1 X 10(-3) M behaved like Ca2+-free incubation but produced sustained contractions with muscle stimulation. Ethanol depressed "off" contractions more than tone and did not affect carbachol-induced contractions. These results suggest that tone probably results from inward leak of Ca2+, whereas "off" contractions depend on release of Ca2+ sequestered in the cell by a mechanism not immediately dependent on increased Ca2+ influx. Carbachol may increase Ca2+ influx as well as utilize sequestered Ca2+. Nifedipine and verapamil may act to block both resting and stimulated Ca2+ influx. Na nitroprusside may act by increasing Ca2+ efflux. Ethanol may act by decreasing the availability of sequestered Ca2+ or by inhibiting the function of a mediator responsible for "off" contractions.     

Effects of nimodipine,Bay K 8644 and pinacidil on α1- and α2-adrenoceptor-mediated vasoconstriction in human hand veins

The effects of nimodipine, Bay K 8644 and pinacidil, three drugs interfering with transmembrane Ca²⁺ fluxes in different ways, were investigated in isolated human hand veins. Their ability to influence the concentration-response relationship for noradrenaline (NA) was assessed in the absence and presence of prazosin or rauwolscine. The contractile response to NA was almost abolished in Ca²⁺-free medium. Nimodipine and pinacidil depressed the NA concentration-response curve both in the absence and presence of α-adrenoceptor blockers. The NA response was only partially inhibited by nimodipine, indicating that NA may activate nimodipine-insensitive influx pathways, presumably receptor-operated calcium channels. Pinacidil inhibited the contractile response to 124 mM K⁺ and reduced the NA-induced contraction in the presence of nimodipine, suggesting that pinacidil has actions other than the opening of potassium channels and subsequent membrane hyperpolarization. Bay K 8644 increased the NA potency fourfold in the presence of rauwolscine, whereas it had no effect on the NA response in the presence of prazosin and in the absence of α-adrenoceptor blockade. Such an action of Bay K 8644 can be reconciled with α-adrenoceptor activation causing membrane depolarization and opening of potential-operated calcium channels. It may be concluded that both α₁- and α₂-adrenoceptor-mediated contractions in human hand veins are highly dependent on Ca²⁺ influx, although the mechanisms utilized to bring about this influx partly differ between the two receptor subtypes.     

Actions of some vasodilators on isolated human hand veins

Vasospasm is a well recognized complication during microvascular surgery of the hand. In the search for new spasmolytic drug therapies, the effects of papaverine, nitroprusside, nimodipine and lidocaine on isolated human hand veins contracted by several postulated mediators of vasospasm were examined. Mechanical activity was recorded isometrically in ring segments of the vessels. Potassium ions, noradrenaline (NA), 5-hydroxytryptamine (5-HT) and prostaglandin F2 alpha (PGF2 alpha) all produced strong contractions that were highly dependent on the presence of extracellular Ca2+. Papaverine acted as a nonselective vasodilator, as it produced an almost identical inhibition of contractile responses to all examined stimulants. Nitroprusside inhibited contractions induced by agonists more than those evoked by K+, whereas the opposite was found for nimodipine. Nitroprusside also seemed to display a certain degree of selectivity among the agonist-induced responses (NA greater than PGF2 alpha greater than 5-HT). Lidocaine increased the contractile response to K+ and at high concentrations (greater than 10(-5) M) produced a contraction per se. The clinical efficacy of lidocaine as a vasodilator after topical application therefore seems to reflect an inhibitory action on vasoconstrictor nerves. Papaverine, nitroprusside, nimodipine and lidocaine differ considerably in their profiles of action and therefore deserve to be further evaluated in the treatment of vasospasm during microvascular surgery.     

Mobilization of intracellular Ca(2+) by thromboxane A(2) does not affect Ca(2+)-activated K(+) channels in rat colonic crypt cells

The effects of 9,11-epithio-11,12-methano-thromboxane A(2) (STA(2)), a stable thromboxane A(2) analogue, and carbachol on colonic Ca(2+)-activated K(+) channels were studied. In indo-1-loaded single cells in isolated rat colonic crypts, both STA(2) (0.1 microM) and carbachol (10 microM) transiently increased intracellular free Ca(2+) concentration ([Ca(2+)](i)) by 136 and 155 nm, respectively. In whole-cell current-clamp experiments of the colonic crypt cells with Cl(-)-free solutions, carbachol (10 microM) hyperpolarized the cell by 19.7 mV, while STA(2) (0.1 microM) did not affect the membrane potential. In the isolated colonic mucosa that was permeabilized mucosally by a monovalent ionophore nystatin in the presence of a serosally directed K(+) gradient, carbachol (10 microM) transiently elicited K(+) current, but STA(2) (0.1 microM) did not. These results indicate that STA(2) elevates [Ca(2+)](i) in rat colonic crypt cells but does not activate basolateral Ca(2+)-activated K(+) channels.     

Influence of extracellular calcium and nifedipine on alpha 1- and alpha 2-adrenoceptor-mediated contractile responses in isolated rat and cat cerebral and mesenteric arteries

The influence of extracellular Ca2+ and nifedipine on contractile responses to 10 microM noradrenaline (NA) was investigated in isolated rat and cat middle cerebral (RCA, CCA) and mesenteric (RMA, CMA) arteries. In the CCA (containing predominantly alpha 2-adrenoceptors), the NA-induced contractions developed considerably more slowly than in the RCA, RMA (containing mainly alpha 1-adrenoceptors) and CMA (sensitive to both alpha 1- and alpha 2-adrenoceptor selective antagonists). The tonic component of the NA-induced contraction in the four types of artery was substantially suppressed after only short periods in Ca2+-free solution. In each type of artery, excluding the CCA, the contractile response to 124 mM K+ was more sensitive to Ca2+ deprivation than that to NA. This suggests that NA, besides mobilizing extracellular Ca2+, can also release Ca2+ from an intracellular pool in the RCA, RMA and CMA, but not in the CCA. Thus, alpha 1-adrenoceptor-mediated contractions in the RCA and RMA seem to depend on both Ca2+ influx and intracellular Ca2+ release, whereas alpha 2-adrenoceptor-mediated contractile responses in the CCA appear to rely almost entirely on Ca2+ influx. Both the maximum response and the tonic component of the NA-induced contraction were significantly more sensitive to nifedipine in the CCA than in the RCA. In comparison with the NA-induced contractions in these arteries, those in the RMA and CMA were relatively resistant to nifedipine. In the CCA exposed to NA in Ca2+-free medium, nifedipine almost abolished the contraction induced by re-addition of Ca2+, whereas in the other types of artery, Ca2+ re-application evoked a significant contraction also in the presence of the drug. The differential effects of nifedipine presumably reflect differences between the arteries, not only in the relative contribution of Ca2+ influx and intracellular Ca2+ release to the contractile activation, but also in the nifedipine sensitivity of the Ca2+ entry pathways utilized by NA. It is concluded that the mechanisms through which NA induces contraction seem to be related both to the subtype of alpha-adrenoceptor stimulated by NA and to the type of vessel studied.     

Characterization of two different calcium entry pathways in small mesenteric arteries from rat

The effects of Ca2+ removal, nifedipine, and La3+ on contractions induced by 124 mM K+ and 10 microM noradrenaline (NA) were investigated in small mesenteric arteries from rat. Ring segments of the arteries were suspended between two steel wires in a 2.5 ml muscle bath, and the mechanical activity recorded "isometrically". The tonic components of the contractile responses to both K+ and NA were critically dependent on the presence of Ca2+ in the bath solution. Nifedipine effectively relaxed K+-contracted arteries, whereas those activated by NA were considerably less affected by the drug. Application of NA to arteries depolarized by K+ in the presence of nifedipine induced a sustained tonic contraction, which was only approximately 20% smaller than that elicited by NA in "standard" Krebs solution, implicating pharmacomechanical coupling. Unlike nifedipine, La3+ inhibited K+- and NA-induced contractions to approximately the same extent. Re-application of Ca2+ to "Ca2+-depleted" preparations exposed to K+ and/or NA induced concentration-dependent contractions. The experimental results suggested that the effects of K+ and NA on the membrane permeability to Ca2+ were additive. The Ca2+-induced contractions were more inhibited by nifedipine in K+-depolarized than in NA-exposed arteries. It is concluded that K+ and NA utilize partly different Ca2+ entry pathways to increase the myoplasmic Ca2+ concentration in rat mesenteric arteries. Whereas K+ seems to promote the influx Ca2+ by activation of CA2+ channels sensitive to the membrane potential, the nature of the receptor-operated Ca2+ entry pathway remains to be established.     

Effect of phorbol 12,13-dibutyrate on smooth muscle tone in rat stomach fundus.

We investigated the effects of phorbol 12,13-dibutyrate (PDBu), a typical protein kinase C (PKC) activator, on smooth muscle tone in the rat stomach fundus. In 5-hydroxytriptamine (5-HT)-precontracted stomach fundus strips, PDBu induced dose-dependent relaxation, but 4alpha-phorbol 12,13-didecanoate, a phorbol ester that does not activate PKC, did not induce relaxation. A PDBu-induced dose-dependent relaxation was also observed in strips precontracted with platelet-activating factor (PAF), carbachol, or 60 mM K+. In stomach fundus strips pretreated with PDBu, the contractile responses to 5-HT and PAF were completely blocked, but those induced by carbachol and endothelin-1 (ET-1) were only partially inhibited. In stomach fundus strips preincubated with carbachol in Ca2+-free medium, the Ca2+-induced contraction was decreased by preincubation with PDBu. In strips preincubated with 5-HT, PAF, or ET-1 in Ca2+-free medium, Ca2+-induced contractions were greatly inhibited by pretreatment with PDBu. These results suggest that in rat stomach fundus strips, PDBu-induced relaxation is mediated by activation of PKC. We speculate that a major factor mediating the relaxant action of PDBu in rat stomach fundus smooth muscle is represented by a reduction in Ca2+ influx via an inhibition of Ca2+ channels.     

Neuromuscular actions of endothelin on smooth, cardiac and skeletal muscle from guinea-pig, rat and rabbit

The peptide endothelin (human, porcine) was investigated for effects on basal muscle tone and on responses to transmural nerve stimulation in a series of smooth muscle preparations, as well as in guinea-pig atrium and rat and guinea-pig diaphragm. Endothelin lacked effect on basal tone or on spontaneous and electrically driven contractions in skeletal and atrial muscle. It contracted guinea-pig ileum, pulmonary and femoral arteries, rat anococcygeus, vas deferens and urinary bladder and rabbit taenia coli, whereas guinea-pig taenia was relaxed. Guinea-pig urinary bladder and vas deferens and rabbit iris sphincter were unaffected up to 3 x 10(-8) M. Endothelin thus has a unique pattern of smooth muscle effects, exhibiting mostly contractile but also relaxing effects. Endothelin modified contractile responses to transmural nerve stimulation, yielding marked and persistent enhancement, in guinea-pig and rat vas deferens, and enhancement also in guinea-pig pulmonary artery. In guinea-pig and rat vas deferens the response to exogenous ATP was increased by endothelin, thus suggesting a strong post-junctional enhancement of neurotransmission. In guinea-pig ileum nerve-induced responses were inhibited by endothelin, whereas exogeneous acetylcholine was enhanced, an effect suggesting a simultaneous pre-junctional inhibition and post-junctional enhancement. The Ca2+ channel blocker felodipine counteracted the stimulatory effects of endothelin on tone and transmurally induced contractions. Tachyphylaxis to endothelin action was sometimes evident, but the anococcygeus being less prone to this might be useful for studies on endothelin antagonism. Endothelin thus has prominent post-junctional, and also probably pre-junctional, effects, lending further support for a distinct biological role of this peptide.     

Differences in contractile activation between human myometrium and intramyometrial arteries

Small intramyometrial arteries and pieces of adjacent myometrial tissue were obtained from 25 non-pregnant women undergoing hysterectomy. Vascular and myometrial preparations were dissected, mounted in organ baths and isometric tension was recorded. Myometrial strips, but no vascular preparation, developed spontaneous contractile activity. Noradrenaline (NA) and vasopressin (VP) contracted both vessels and myometrium. Prostaglandin F_2α (PGF_2α) contracted the myometrial tissue, but had only a minor effect on the vessels. Removal of extracellular calcium almost abolished the myometrial responses to high K⁺ (124 mm)-solution, PGF_2α, NA and VP. The vascular responses remaining after this treatment were 18% (K⁺), 34% (NA) and 25% (VP) of control contractions induced by high K⁺ (124 mm). Nifedipine potently depressed myometrial contractions induced by NA and VP, but was less active against the vascular responses to these agents. In preparations exposed to calcium-free medium, nifedipine (10^-7 m) almost abolished myometrial contractions induced by calcium in the presence of K⁺ (124 mm), NA or VP. It also effectively depressed vascular responses to calcium in the presence of K⁺, but was less active if NA and VP were present. It is suggested that PGF_2α has almost no contractant effect on intramyometrial arteries, and that the activation process in these vessels is much less dependent on extracellular calcium than that of the myometrium.     

Cyclic AMP-mediated inhibition of noradrenaline-induced contraction and Ca2+ influx in guinea-pig vas deferens

The relaxation effects of forskolin and methylxanthines on noradrenaline (NA)-induced contractions were investigated by measuring isotonic contraction and intracellular calcium concentration ([Ca2+]i) in the epididymal side of guinea-pig vas deferens. NA (100 microM) and high K+ (55 mM) induced a biphasic contraction; fast, transient (phasic) and slow, sustained (tonic) phases. Both phases in either NA or high K+ stimulation were abolished in Ca2+-free solution. Pretreatment with 10 microM nifedipine, an L-type Ca2+ channel blocker, reduced both phasic and tonic contractions induced by high K+. In the case of NA-induced contraction, however, nifedipine reduced the phasic contraction but not the tonic contraction. The nifedipine-insensitive tonic contraction was relaxed by the application of polyvalent cations (Mn2+, Co2+, Cd2+ and La3+). These findings indicate that NA-induced biphasic contraction is mainly due to nifedipine-insensitive Ca2+ influx, especially in the tonic phase. Cyclic AMP-increasing agents such as forskolin (0.5-10 microM), IBMX (5-500 microM) and caffeine (1-20 mM) relaxed the NA-induced contraction extensively in a concentration-dependent manner. However, these agents only partially relaxed the high K+-induced contraction. Forskolin (10 microM) and IBMX (100 microM) reduced the [Ca2+]i response to NA, but had no effect on the [Ca2+]i response to high K+. These results suggest that an increase in intracellular cAMP may relax the NA-induced contraction by attenuating a nifedipine-insensitive Ca2+ influx and by a mechanism independent of a reduction in [Ca2+]i.     

Distribution and function of nitric oxide-containing nerves in canine corpus cavernosum and spongiosum

Specimens of penile erectile tissue from the corpus cavernosum (CC) and corpus spongiosum (CS) of beagle dogs were investigated with reference to morphological and functional aspects of the nitric oxide (NO) system. Using immunohistochemistry, the smooth muscle bundles of the CC were found to contain a rich innervation, as seen by the pan-neuronal marker, protein gene product 9.5. A large number of nerves also stained positively for acetylcholine esterase. There was a moderate to rich supply of nerves containing NO synthase, which was frequently co-localized with vasoactive intestinal polypeptide. CS preparations from the proximal penis (the spongious bulb) had an innervation similar to that of the CC. However, CS preparations from the distal penis had a distinctly lower density of nerves than the proximal part, and spongious tissue from the glans penis was practically devoid of nerves. In isolated preparations of proximal and distal CC and CS, contracted by noradrenaline, electrical field stimulation (EFS) produced frequency-dependent relaxations. The responses were less pronounced in preparations from the distal penis, particularly in the CS. Pre-treatment with the NO-synthesis inhibitor N^G-nitro-L-arginine abolished all relaxations. Carbachol effectively relaxed noradrenaline-contracted strips. The responses were similar in proximal preparations from the CC and CS, and significantly less pronounced in strips from the distal penis. N^G-nitro-L-arginine significantly reduced the relaxant effect of carbachol. Preparations of spongious tissue from the glans penis did not react to high potassium solution, addition of noradrenaline, or EFS. The results demonstrate regional differences within the CS, both in innervation and functional responses.     

Source of calcium for contractions induced by depolarization and muscarinic receptor stimulation in rabbit urinary bladder

Omission of calcium or the inclusion of lanthanum in the bathing medium resulted in an almost complete inhibition of contractile responses induced by either K+ depolarization or carbachol in strips of rabbit urinary bladder. D-600 inhibited K+-induced contractions significantly more than carbachol-induced responses. The influx of 45Ca into cells was stimulated both by K+ depolarization and carbachol. Over a 2-min period the increase in 45Ca influx induced by high K+ and carbachol was 98 and 65%, respectively. Both lanthanum and D-600 blocked 45Ca influx stimulated by either K+ depolarization or carbachol. The inhibition of 45Ca influx by these calcium-channel blocking agents, particularly by D-600, was dependent on the length of exposure. Application of carbachol during 45Ca efflux in pre-loaded muscle strips had no effect on the rate of 45Ca efflux. These results indicate that the contractile responses of the urinary bladder to depolarization and to carbachol are highly dependent on an extracellular source of calcium.