Segments Of Rat Vas Deferens Research Articles

Some pharmacological studies of venom from the inland taipan ( Oxyuranus microlepidotus)

The present study was designed to obtain a basic pharmacological profile of venom from the inland taipan ( Oxyuranus microlepidotus). Venom (0.05–50 μg/ml) produced dose-dependent contractions in guinea-pig ileum, which could not be reproduced upon second administration. The cyclooxygenase inhibitor indomethacin (1 μM), a preceding anaphylactic response induced by egg albumin and inactivation of phospholipase A 2 (PLA 2) by incubation with 4-bromophenacyl bromide (1.8 mM) all significantly inhibited responses to venom (0.5 μg/ml). Venom (0.5 μg/ml) caused inhibition of stimulation-induced contractions in the prostatic segment of rat vas deferens which was not significantly affected by the α 2-adrenoceptor antagonist idazoxan (0.3 μM). Venom (10 μg/ml) caused time-dependent inhibition of the rat electrically stimulated phrenic nerve–diaphragm preparation, positive inotropic and chronotropic responses in rat isolated atria and relaxation in rat endothelium-denuded and -intact isolated aortae. In endothelium-intact aortae, the nitric oxide synthase inhibitor N-nitro- l-arginine (NOLA, 0.1 mM) significantly inhibited the response to venom (10 μg/ml). Venom (50 μg/kg, i.v.) caused an immediate drop in blood pressure followed by cardiovascular collapse in anaesthetised rats. Venom (10 μg/kg, i.v.) caused a gradual fall in blood pressure which was sometimes accompanied by a temporary cessation of respiration. A PLA 2 assay detected the presence of PLA 2 in the venom. These results suggest that the venom contains a component capable of causing the synthesis of arachidonic acid metabolites and a component capable of relaxing vascular smooth muscle. The inhibitory effect on the phrenic nerve–diaphragm is probably due to the previously identified neurotoxin (paradoxin).

Evidence for adrenergic and tachykinin activity in venom of the stonefish ( Synanceja trachynis)

The aim of the present study was to investigate previously suggested adrenergic and tachykinin activity, as well as the cardiovascular effects, of venom from the stonefish (Synanceja trachynis). Stonefish venom (60-120 micrograms/kg, i.v.) produced dose-dependent bronchoconstriction in anaesthetised guinea-pigs. This response (100 micrograms/kg, i.v.) was significantly reduced by the neurokinin 1 (NK1) receptor antagonist CP-99,994 (1 mg/kg, i.v.). Contractile responses to venom (4 micrograms/ml) of guinea-pig isolated ileum (GPI) were significantly inhibited by a combination of the sodium channel blocking drug tetrodotoxin (1 microM) and the ganglion blocking drug mecamylamine (10 microM). However, subsequent administration of CP-99,994 (0.1 microM) did not produce further inhibition. Endogenous tachykinin depletion with capsaicin (1 microM) also significantly attenuated responses to venom (4 micrograms/ml) in GPI. Venom (4 micrograms/ml) produced increases in rate and force of contraction of rat spontaneously beating isolated atria which were significantly inhibited by the beta-adrenoceptor antagonist propranolol (5 microM) but not by noradrenergic transmitter depletion with reserpine (4.5 mg/kg, i.p.). In the presence of the alpha 1-adrenoceptor antagonist prazosin (0.3 microM), venom (6 micrograms/ml) significantly inhibited electrically evoked twitches of prostatic segments of rat vas deferens. The inhibitory effect of venom was significantly reduced by the alpha 2-adrenoceptor antagonist idazoxan (1 microM) but not by propranolol (5 microM) or the neurokinin 2 (NK2) receptor antagonist SR-48,968 (0.1 microM). Venom (60-120 micrograms/kg, i.v.) produced dose-dependent increases in mean arterial blood pressure in anaesthetised rats. This pressor response (60 micrograms/kg, i.v.) was significantly reduced by prazosin (10-50 micrograms/kg, i.v.) and the leukotriene receptor antagonist SB205312 (1 mg/kg, i.v.), significantly increased by propranolol (2 mg/kg, i.v.), but not significantly affected by the cyclo-oxygenase inhibitor indomethacin (10 mg/kg, i.v.) or the thromboxane A2/prostaglandin H2 (TP) receptor antagonist GR32191B (1 mg/kg, i.v.). Pressor responses to venom (100 micrograms/kg, i.v.) were also observed in anaesthetised rabbits. These results suggest that stonefish venom contains a component capable of stimulating the release of endogenous tachykinins with subsequent activity at NK1 receptors. The venom also appears to act via stimulation of sodium channels on sensory nerves. The venom also has activity at alpha 2-adrenoceptors and a direct action at beta-adrenoceptors. The effect of venom on blood pressure of anaesthetised rats appears to include a pressor component that is mediated, in part,by alpha-adrenoceptors and leukotriene receptors, and a depressor component that is mediated by beta-adrenoceptors. However, the pressor response does not involve action at TP receptors, or require the production of cyclo-oxygenase metabolites.

Two phases of contractile response in rat isolated vas deferens and their regulation by adenosine and α-receptors

Contractions to transmural electrical stimulation and exogenous norepinephrine were recorded in isolated longitudinal segments of rat vas deferens. Electrical stimulation for 30 s produced a biphasic contraction in the vas deferens consisting of a rapid, transient response (Phase I), followed by a slowly developing, sustained contraction (Phase II). N 6-Cyclohexyladenosine (CHA), a selective adenosine 1 (A 1)-receptor agonist, attenuated in a concentration-dependent manner the Phase I contractile response, while having little effect on the Phase II response. In contrast, 2-(phenylamino)adenosine (CV-1808), a selective adenosine 2 (A 2)-receptor agonist had little effect on either contractile phase. CHA did not inhibit the contraction to exogenous norepinephrine, suggesting that A 1-receptors were located at a presynaptic site. The relatively selective α 2-receptor agonist clonidine produced the same pattern of contractile inhibition as CHA. The inhibitory effect of CHA on the Phase I contractile response in the vas deferens could be antagonized by the selective A 1-receptor antagonist 8-cyclopentyltheophylline, while the selective α 2-receptor antagonist idazoxan preferentially antagonized the inhibitory effect of clonidine on the Phase I response. Both the Phase I and Phase II contractile responses were reduced by the selective α 1-adrenoceptor antagonist prazosin and the ATP analog α,β-methylene adenosine triphosphate (α,β-methylene ATP), suggesting that norepinephrine and ATP are coreleased as neurotransmitters for both responses. The results of the present study demonstrate that in the rat vas deferens the presynaptic inhibitory effects of adenosine is mediated by the A 1-receptor subtype, and that both A 1- and α 2-receptor agonists exert a selective inhibitory effect on the Phase I contractile response to electrical stimulation.

A study of stimulation-evoked activation of alpha 2-adrenoceptors in the rat isolated vas deferens.

Experiments were performed with prostatic and epididymal segments of rat vas deferens to determine whether alpha 2-adrenoceptors in this organ were activated during field stimulation of sympathetic terminals with short trains of pulses applied at low frequencies. In prostatic segments the magnitude of twitches evoked by trains of ten field pulses at 1 or 2 Hz declined after 2-3 s of stimulation. In contrast, facilitation of twitches and fusion of contractions occurred when similar stimulation was applied to epididymal segments. In prostatic segments from rats treated with reserpine (2.5 mg/kg i.p.) 24 h previously, there was no decline in the magnitude of twitches produced by successive impulses in a train of stimulating pulses. In epididymal segments from reserpine-treated rats facilitation of twitches in response to successive impulses in each train still occurred. In prostatic segments cocaine (5 and 10 mumol/l) enhanced twitch fade with stimulation at 1 and 2 Hz without altering the time for onset of this effect. In epididymal segments cocaine led to enhancement and prolongation of contractile responses. In prostatic segments yohimbine (0.01-0.06 mumol/l) reduced or reversed the effect of cocaine in enhancing twitch fade. In preparations where the reversal of the effect of cocaine by yohimbine was incomplete, subsequent addition of phentolamine (1 mumol/l) produced complete reversal. In epididymal segments yohimbine (0.01 mumol/l) produced a further enhancement in the twitch responses to stimulation at 1 and 2 Hz. Subsequent addition of phentolamine (1 mumol/l) reversed the facilitatory effects of cocaine and yohimbine. Propranolol (10 mumol/l) was without effect upon responses to stimulation in either segment of rat vasa deferentia. These experiments indicate that noradrenaline, released by short trains of impulses applied at low frequency to hypogastric nerve terminals activates prejunctional alpha 2-adrenoceptors in the prostatic segment of the vas deferens. In the epididymal portion the effects arising from activation of prejunctional alpha 2-adrenoceptors are outweighed by the consequences of activation of extrajunctional alpha 1-adrenoceptors located on longitudinally arranged muscle.

Increases in cyclic GMP levels may not mediate relaxant effects of sodium nitroprusside, verapamil and hydralazine in rat vas deferens

IT has been suggested that increases in cyclic GMP levels may be responsible for the promotion of contractions in a variety of smooth muscles1–6. This suggestion was based on observations that agents known to be capable of contracting smooth muscles produced marked increases in cyclic GMP levels in the respective muscles1–8. However, recent reports have concluded that, although increases in cyclic GMP levels do occur during contractions in some types of smooth muscle, the increases in cyclic GMP levels are not responsible for the initiation or promotion of the contractions9–12. Part of the evidence for this conclusion was the observation that marked increases in cyclic GMP levels occurred during relaxation of smooth muscles by drugs such as nitroglycerol10,12. Schultz et al.13 have shown that other smooth muscle relaxants including sodium nitroprusside, hydralazine, and D-600 (a methoxy derivative of verapamil) are all capable of increasing cyclic GMP levels in isolated segments of rat vas deferens. These authors also noted that the 8-bromo derivative of cyclic GMP was itself capable of relaxing the rat vas deferens in certain conditions13. It was concluded that, not only was cyclic GMP not a mediator of smooth muscle contraction, but it might in fact be responsible for the smooth muscle relaxant effects of the above drugs. We describe here studies performed in rat vas deferens. Our results are not consistent with the hypothesis that increases in cyclic GMP levels are responsible for drug-induced smooth muscle relaxation.

Effects of magnesium ions on adrenergic nerve terminals.

Desheathed segments of rat vas deferens were incubated in modified Krebs solutions containing 10, 40, or 80 mM MgCl2 for periods ranging from 10 to 90 min. The tissue specimens were prepared for fluorescence histochemistry and electron microscopy. The fluorescence of the incubated specimens was comparable to that of fresh-fixed and Krebs-incubated controls. Ultrastructural observations showed that longer incubations and higher concentrations of Mg2+ were correlated with the increased tendency for synaptic vesicles to adhere to the axonal plasmalemma forming five-layered junctions. Alternatively, strings of vesicles were observed within delicate spike-like projections of the axons. In addition, clusters of vesicles or single vesicles were often observed periaxonally.