Effects of streptozotocin-induced diabetes and insulin on calcium responsiveness of the rat vas deferens

In the present study, effects of streptozotocin-induced diabetes and insulin treatment on the reactivity of rat vas deferens to KCl and calmidazolium, a calmodulin antagonist, were evaluated and calmodulin levels in vas deferens tissue from diabetic and insulin-treated rats were determined. Diabetes was induced in rats by a single injection of streptozotocin. Five weeks after the induction of diabetes, one group of diabetic rats was injected with insulin for 3 weeks. After 8 weeks, vas deferens tissues on one side of diabetic and insulin-treated diabetic rats and their controls were mounted in organ bath to measure isometric tension, while the tissues on the other side of rats were homogenized to determine calmodulin levels by radioimmunoassay. Concentration-response curves to KCl were obtained in vas deferens tissues in the absence and presence of calmidazolium. The effects of KCl and calmidazolium on vas deferens isolated from 8-weeks diabetic rats were decreased. Calmodulin levels were also found to be decreased in vas deferens from diabetic rats. Decreased calmodulin levels in diabetic rat vas deferens were not corrected by insulin treatment. Only a partial correction following insulin treatment was observed in contractile effect of KCl on diabetic rat vas deferens, whereas insulin treatment increases the affinity of calmodulin in this muscle. Experimental diabetes causes an impairment in calcium/calmodulin-dependent contractile process of vas deferens, which is correctable partially following insulin therapy. The changes in the function of rat vas deferens due to streptozotocin diabetes seem to be related to impaired sexual functions in human diabetes.     

Influence of repeated oscillation stress in rats

The influence of two weeks of repeated oscillation stress on rats was investigated. Organ weights, liver glycogen, adrenal ascorbic acid, hematological and serum biochemical analysis and maximum contraction of vas deferens induced by noradrenaline were measured. In stress-loaded rats, body weight fell to 85-90% compared with control rats. Atrophy of the thymus and hypertrophy of the adrenals were found in stress-loaded rats. Hematological and serum biochemical findings revealed that white blood cells and blood glucose decreased, but NEFA increased significantly. Serum sodium increased, but potassium decreased. Maximum contraction of vas deferens induced by noradrenaline was potentiated in stress-loaded rats. The other findings did not differ from those of controls. From these results, it is suggested that the stress-loaded rat shows some abnormalities, but may adapt partially to stress.     

Morphine-like activity of the enkephalin analog Tyr-D-Ala-Gly-Phe-NH2

The analgetic activity of the tetrapeptide enkephalin analog, its influence on the interneuronal transmission of excitation in various areas of the central nervous system and on opiate receptors of vas deferens were studied. The tetrapeptide was found to have a marked analgetic effect during intravenous injection to mice but to be less active than morphine. The tetrapeptide as well as morphine inhibited the impulse summation in rabbits and both spontaneous and bradykinin-induced neuronal activity in the rat sensory motor cortex. The tetrapeptide inhibited the contractions of isolated vas deferens in mice. The opiate antagonist naloxone eliminated both analgetic effect of the tetrapeptide and its inhibitory effect on the impulse summation, neuronal activity and contractions of vas deferens.     

Spontaneously hypertensive rats exhibit an enhanced mydriatic response to clonidine. Evidence for enhanced sensitivity of central alpha 2-adrenoceptors

Comparisons among spontaneously hypertensive (SHR), Kyoto Wistar (KW), and Wistar (W) rats were made of the functional states of central nervous system (CNS) alpha 2-adrenoceptors (clonidine-induced mydriasis) and nonvascular peripheral presynaptic alpha 2-adrenoceptors (clonidine-induced inhibition of the neurogenic twitch of the isolated vas deferens). While there were no differences among the strains of rats in the concentration of clonidine required to produce a 50% inhibition of the electrically evoked contractile response of the vas deferens, there was a significant reduction in the mean effective concentration (ED50) of clonidine to induce mydriasis in SHR as compared with KW and W rats. These observations indicate that CNS alpha 2-adrenoceptors may be functionally more sensitive in SHR. The data also suggest that the sensitivity of nonvascular presynaptic alpha 2-adrenoceptors, at least in the vas deferens, is not altered in hypertensive animals.     

Action of pro-opiomelanocortin products on the rat vas deferens

Behavioral study of pro-opiomelanocortin products indicates that beta-endorphin and corticotrophin-like peptides have antagonistic effects. However, these peptides have similar actions on the rat vas deferens. beta-endorphin, alpha-MSH and ACTH each inhibit electrically evoked contraction of the duct, but the corticotrophin derived peptides are tenfold more potent on a molar basis (ED50 = 9 nM). Pharmacological analysis shows that the action of corticotrophin-derived peptides does not involve an opiate receptor mechanism. The results are discussed in terms of the central action of the peptides.     

A pharmacological investigation of the biphasic nature of the contractile response of rabbit and rat vas deferens to field stimulation

It has been demonstrated previously with the vas deferens of the guinea-pig that the first and second phases of the contractile response to motor nerve stimulation are preferentially antagonized by the P₂-purinoceptor antagonist arylazido aminopropionyl ATP (ANAPP₃), and the α₁-adrenoceptor antagonist prazosin, respectively. We have now investigated the effect of the two antagonists on the biphasic contraction in the vas deferens of two other species; rabbit and rat. ANAPP₃, in a concentration which antagonized responses to exogenously applied ATP but not those to exogenous norepinephrine, preferentially reduced the initial phasic response of the rabbit vas deferens to motor nerve stimulation without significantly reducing the secondary, tonic phase of the response. Prazosin had the opposite effect; antagonizing the response to norepinephrine but not to ATP and reducing the tonic response to motor nerve stimulation without significantly reducing the initial phasic response. Results obtained with the rat vas deferens were similar. The present results combined with previous findings suggest that ATP and norepinephrine act as cotransmitters in the vas deferens of several species.     

The effect of cocaine on kinetics of α<Subscript>1</Subscript>-adrenergic contractile response in different portions of the rat vas deferens

The effect of cocaine (10 μM) on the kinetics of contractile response to noradrenaline (NA) was studied in the rat epididymal and prostatic vas deferens. Cocaine caused an acute increase in vas deferens adrenergic sensitivity primarily due to blockage of NA neuronal capture. The presynaptic action prevailed in the epididymal half: the EC 50 value decreased 32-fold with a slight increase of the maximum adrenergic response more evident in the prostatic half. In the presence of cocaine, the prostatic contraction to NA was mediated not by a single pool of α₁-adrenoceptors like in epididymal vas deferens but by two. Its high affinity pool had the same affinity as α₁-adrenoceptors of the epididymal half, the affinity value of the low one was 36-fold less, and the total maximal response of both pools increased 4.5-fold. The differences in cocaine effect on the rat epididymal and prostatic vas deferens contractions to NA appear to be caused by the different sources of Ca²⁺ involved in these responses.     

The androgenic effect of norethisterone and 5alpha-norethisterone on the contractile response of the rat vas deferens to methoxamine and serotonin.

Norethisterone (NET) and its metabolite 5alpha-norethisterone (5alpha-NET) are competitors for the androgen receptor. The sensitivity of the rat vas deferens to the contractile action of methoxamine and serotonin is regulated by hormonal and anatomical factors. The aim of this study was to evaluate the ability of NET and 5alpha-NET to induce the androgen-regulated contractile response to methoxamine and serotonin in the epididymal and prostatic portions of rat vas deferens. Adult male rats either intact, castrated or steroid-treated castrated were used. The contractility was recorded isometrically, and non-cumulative concentration-response curves to either methoxamine or serotonin were obtained. NET and 5alpha-NET partially restored the sensitivity to methoxamine and serotonin in the epididymal portion of castrated rats. The maximal responses to both agonists were significantly higher than those observed in castrated rats, and significantly lower than the responses observed in either intact or androgen-treated castrated rats. The prostatic portion was less responsive to both agonists than the epididymal portion, in all groups but castrated rats, as castration induced sensitivity to both agonists. NET and 5alpha-NET displayed a partial though similar androgenic activity in the rat vas deferens. These results contrast with previous reports where a decrease of androgenic effect due to the 5alpha-reduction of NET has been found.     

Development of tolerance to the excitatory effect of morphine and cross-tolerance to the inhibitory action of β-endorphin in the isolated rat vas deferens

In the isolated rat vas deferens, morphine caused an increase in the neuromuscular twitch evoked by electrical stimulation. In rats chronically treated with morphine, the concentration of the opiate required to cause a 50% increase in the twitch was about 3 times larger than needed to elicit the same response in vasa from rats treated with a placebo. The simultaneous administration of naloxone plus morphine partially antagonized tolerance development by about 22%. Morphine tolerance was extended to other opiate- like alkaloids such as etonitazene and a derivative of azidomorphine (CAM). In contrast to the effects of morphine, β-endorphin inhibited neuromuscular transmission. Vasa from rats chronically treated with morphine were about 10 times less sensitive to the inhibitory effect of β-endorphin as compared to paired placebo treated controls. Chronic naloxone treatment in conjuction with morphine significantly reduced the cross-opiate tolerance by 66%. Present results suggest that morphine may interact at two different sites in the nerve terminals of the rat vas deferens.     

Development of clonidine-tolerance in the rat vas deferens: Cross tolerance to other presynaptic inhibitory agents

Possibility of the development of clonidine-tolerance in the peripheral nervous tissue was examined using vas deferens isolated from rats chronically treated with clonidine. Rats were treated with clonidine for 10 days by adding the drug to drinking water (10 μg/ml). For the control rats, drug-free tap water was provided. Electrically evoked twitch response of vas deferens was suppressed by adenosine, β-endorphine and α₂-adrenergic agonists, such as clonidine and B-HT 933, both in control and clonidine-treated groups. Vas deferens isolated from clonidine-treated rats showed significantly lower responsiveness to the inhibitory effects of clonidine and B-HT 933 compared to those from control rats. Vas deferens from clonidine-treated rats also was less responsive to adenosine and β-endorphin, both of which interact with presynaptic inhibitory receptors other than α₂-adrenergic and muscarinic cholinergic stimulation responsiveness of the postsynaptic smooth muscle to both α-adrenergic and muscarinic cholinergic stimulation did not change after 10 days of treatment with clonidine. These results suggest that clonidinetolerance can be induced in the peripheral nervous system by chronic treatment of this drug and that the tolerance is not specific to α₂-adrenergic agonists. Some common pathway in the inhibitory mechanisms of various agents or possible interactions between the different types of presynaptic inhibitory receptors may be involved in this phenomenon.     

Effects of veratrine and paeoniflorin on isolated mouse vas deferens

In this study, we attempted to identify the interactions and mechanisms between veratrine and paeoniflorin on isolated mouse vas deferens. Paeoniflorin had no effect on isolated mouse vas deferens. Veratrine (1 x 10(-5) approximately 1 x 10(-3) g/ml) could directly induce contraction of isolated rat and mouse vas deferens. The concentration induced by veratrine (1 x 10(-5) g/ml) was completely inhibited by Ca2+-free solution and verapamil (1 x 10(-5) M), in both the epididymal and the prostatic portions of isolated mouse vas deferens. Naloxone (1 x 10(-5) M) did not alter the contraction induced by veratrine (1 x 10(-5) g/ml) in either the epididymal or the prostatic portions of isolated mouse vas deferens. Paeoniflorin (4.8 x 10(-5) g/ml) inhibited the contraction induced by veratrine (1 x 10(-5) g/ml) in both the epididymal and the prostatic portions of isolated mouse vas deferens. Paeoniflorin (4.8 x 10(-5) g/ml) potentiated norepinephrine (1 x 10(-5) M)-induced phasic contraction in the epididymal portion, but decreased contractions in the prostatic portion. Paeoniflorin (4.8 x 10(-5) g/ml) increased KCI (56 mM)-induced phasic contraction in the epididymal portion, but decreased the tonic contraction in either the epididymal or the prostatic portion. Veratrine (1 x 10(-5) g/ml)-induced contractions could be decreased by pretreatment with ryanodine (1 x 10(-5) M) in both the epididymal and the prostatic portions. Pretreatment with the combination of paeoniflorin (4.8 x 10(-5) g/ml) and ryanodine (1 x 10(-5) M) did not potentiate the inhibition of paeoniflorin in the veratrine-induced contraction in both the epididymal and the prostatic portions of isolated mouse vas deferens.     

Affinity profile of the novel muscarinic antagonist, guanylpirenzepine

The study reports the functional affinity of an amidino derivative of pirenzepine, guanylpirenzepine, for muscarinic receptors mediating relaxation of rat duodenum, inhibition of rabbit vas deferens twitch contraction (both receptors previously classified as M1), guinea pig negative inotropism (M2) and ileal contraction (M3). Unlike pirenzepine, guanylpirenzepine discriminated between duodenum and vas deferens receptors, with a 30-fold greater affinity for the former subtype. The unique selectivity pattern of guanylpirenzepine (duodenum greater than vas deferens greater than ileum greater than atrium) renders it a promising tool for the classification of muscarinic receptor subtypes.     

The effect of sympathomimetic drugs on contractility of the vas deferens in vitro and in vivo.

The sympathomimetic drugs noradrenaline, methoxamine, tyramine and norephedrine caused rhythmic contractions in isolated human vasa deferentia. Provided the drug was not washed out, these contractions lasted for the entire duration of the experiment (4-6 h). These contractions were mediated via alpha-adrenoreceptors. Intravenous administration of methoxamine or oxymetazolene to rats or guinea-pigs produced contractions of the vas deferens in vivo in some experiments but was accompanied by severe cardiovascular side effects. A local method of application was developed, using mixtures of tyramine with Silastic prepared as collars specially designed to fit round the vas deferens. Acute and chronic insertion of these slow-releasing devices around the vas deferens of rats produced rhythmic contractions of the vas deferens without any serious side effects.     

Leu9 psi(CH2NH)Leu10]-neurokinin A (4-10) (MDL 28,564) distinguishes tissue tachykinin peptide NK2 receptors

The neurokinin A analogue, MDL 28,564 (Asp-Ser-Phe-Val-Gly-Leu-CH2NH-Leu-NH2), inhibited 125I-NKA binding to hamster urinary bladder NK2 receptors with a KI of 130 nM. For rat submaxillary gland NK1 receptors and cerebral cortical NK3 receptors, the KI's for MDL 28,564 were greater than 250 microM and greater than 500 microM, respectively. MDL 28,564 did not relax dog carotid artery (NK1 tissue) or contract rat portal vein (NK3 tissue). In guinea-pig trachea tissues, MDL 28,564 stimulated phosphatidylinositol turnover and induced contraction with maximum effects similar to those of neurokinin A. In hamster urinary bladder tissue, MDL 28,564 stimulated phosphatidylinositol turnover with maximum effect only 10% of that of neurokinin A, did not produce sustained contraction itself and antagonized NKA-induced contraction. MDL 28,564 also produced full contraction in rabbit pulmonary artery (NK2 tissue) but was inactive in rat vas deferens (NK2 tissue). These data with MDL 28,564 are consistent with the NK2 receptors in guinea-pig trachea and rabbit pulmonary artery being different from those in hamster urinary bladder and rat vas deferens.     

The effect of hypertonic solution on the wet weight and contractions of rat uterus and vas deferens

The role of propagated activity in the responses to agonist drugs was studied for the rat uterus and vas deferens. Hypertonic solutions were used to inhibit propagation of activity by shrinking cells. Tissue weight was used to indicate cell volume. Hypertonic solutions after 10 min caused weight loss and reduced the size of contractions in response to submaximal doses of drugs, to KCl, and to external electrical stimulation. Contractions in response to KCl and drugs were diminished to a similar degree in the vas deferens, but in the uterus, drug contractions were depressed much more. Prolonged action of hypertonic solution also differed for the two tissues. In the uterus, weight changes correlated with changes in size of the drug-induced contractions. Uterine contractions reduced in hypertonic solution could be increased by using supramaximal doses of drug. When stimulation was applied to one end of the uterus in a three compartment bath, propagation of spontaneous drug- and KCl-induced contraction occurred, but it was prevented by placing hypertonic solution in the center compartment. An increase of the KCl to 44 mM in the hypertonic solution restored propagation. These experiments yielded no evidence of propagated responses in the rat vas deferens. It was concluded that propagated activity plays a role in drug-induced contractions in the rat uterus but not in the rat vas deferens. Hyperpolarization of shrunken cells might be involved in inhibition of propagation by hypertonic solutions.     

Differential contractile response of normal vas deferens of rodents in correlation to their calcium and electrolyte levels

The contractile pattern of the vas deferens in three different rodents, rat, guinea pig and mouse was studied in response to adrenaline and noradrenaline. The left vas deferens of rat was more responsive to the graded doses of adrenaline and noradrenaline than the right. The same was also true for guinea pig and mouse vas deferens. This differential response has been correlated with the greater concentrations of calcium and sodium in the right vas deferens in rats and guinea pigs and it might also be related to the levels of membrane-bound and intracellular calmodulin-bound calcium. It is suggested that the left vas deferens might possess more calmodulin-bound calcium than the right, which might have instead, more membrane-bound calcium.     

Release of contractile agents from rat vas deferens by clonidine.

Clonidine induces contractile effects on the isolated rat vas deferens, but not on rat uterus or guinea-pig ileum. However, we have observed that if clonidine is incubated for about 10 min with a nutrient solution containing an isolated rat vas deferens, the resulting solution can contract an isolated rat uterus, or guinea-pig ileum indicating the involvement of a substance released from the vas. This contractile effect was partially reduced by naloxone and by serotonin antagonists, and by using a denervated vas, indicating that opioids, serotonin and eventually other substances released from nerve tissue of the vas can be involved.     

Testosterone stimulates prostanoid production by rat vas deferens.

The rat isolated vas deferens produces and releases prostanoids into an incubation medium. Production of these substances from the exogenous precursor 14C arachidonic acid was studied in prepubertal, pubertal and adult animals. Synthesis of prostaglandin F, prostaglandin E, prostaglandin D and thromboxane B2 is lower in prepubertals arid increases significantly in pubertals, with no further modifications in adults. Castration of pubertals and adults dramatically reduces the production of all measured arachidonic acid metabolites but does not modify it in prepubertals. Replacement therapy with testosterone propionate significantly enhances prostanoid production in pubertal and adult castrated rats. Similar treatment on normal prepubertals also increases synthesis, indicating that androgens could be modulators of prostanoid synthesis in vas deferens. The lower effects obtained treating castrated adults with progesterone and with 17-beta estradiol suggest an action, at least partially specific for androgenic steroids. It is concluded that prostanoid production by the rat vas deferens from an exogenous precursor is closely related to the presence of androgens.