Evidence that (-)-7-hydroxy-4'-dimethylheptyl-cannabidiol activates a non-CB(1), non-CB(2), non-TRPV1 target in the mouse vas deferens

Previous experiments showed that R-(+)-WIN55212-induced inhibition of electrically-evoked contractions of mouse vasa deferentia could be antagonized by cannabidiol in a manner that appeared to be competitive but not to involve direct competition for established cannabinoid receptors. We have now discovered that (−)-7-hydroxy-4′-dimethylheptyl-cannabidiol (7-OH-DMH-CBD) inhibits electrically-evoked contractions of the vas deferens (EC₅₀ = 13.3 nM). This it appeared to do by acting on prejunctional neurones as 100 nM 7-OH-DMH-CBD did not attenuate contractile responses to phenylephrine or β,γ-methylene-ATP. Although 7-OH-DMH-CBD was antagonized by SR141716A, it was less susceptible to antagonism by this CB₁ receptor antagonist than R-(+)-WIN55212. 7-OH-DMH-CBD was also antagonized by cannabidiol (1 μM; apparent K_B = 222.2 nM) but not by the CB₂ receptor antagonist, SR144528 (32 nM), or by naloxone (300 nM), ruthenium red (1 μM) or capsazepine (10 μM). Yohimbine (100 nM) enhanced the ability of 7-OH-DMH-CBD to inhibit electrically-evoked contractions. R-(+)-WIN55212 was also potentiated by 100 nM yohimbine, possibly reflecting ongoing sequestration of G_i/o proteins from CB₁ receptors by ₂-adrenoceptors. Our results suggest that 7-OH-DMH-CBD may activate a neuronal target in the vas deferens that is not a CB₁, CB₂, TRPV1, opioid or ₂-adrenergic receptor but do not exclude the possibility that it also activates CB₁ receptors.     

Regional Differences in Steroidogenesis and Hormone Levels in the Epididymis and Vas Deferens of Adult Rats

In vivo and in vitro studies with different parts of the epididymis and vas deferens were carried out to determine their inherent capacity to synthesize steroids and to correlate with the endogenous levels with or without the administration of hCG.
Incubation with ¹⁴C-labelled pregnenolone and testosterone demonstrated that caput epididymidis was more active than other parts in synthesizing testosterone from ¹⁴C-pregnenolone and in converting labelled testosterone to 5α-dihydrotestosterone (DHT). The cauda epididymidis and vas deferens accumulated more radioactivity in progesterone and dehydroepiandrosterone (DHEA) than the caput epididymidis.
The levels of DHT, testosterone and 4-androstene-3,17-dione in the caput epididymidis were reduced after ligation of ipselateral efferent ductules indicating the testicular origin of these steroids. The cauda epididymidis and vas deferens had higher levels of progesterone as compared to the other regions of the epididymis, which were decreased after the ligation. Intravenous injection of hCG increased the levels of oestradiol-17β in all tissues and markedly in the cauda epididymidis and vas deferens. The high levels of progesterone and oestradiol-17β present in these organs may be of importance in maintaining fertilizing ability of spermatozoa stored in the cauda epididymidis and vas deferens and their transport.     

KCL CONTRACTIONS IN THE RAT INTACT AND BISECTED VAS DEFERENS: CONTRIBUTION OF ENDOGENOUS NORADRENALINE RELEASE

1 KCl produced a biphasic contraction in the intact rat vas deferens. Both components were larger and the initial rapid phasic component was faster in the prostatic portion than the epididymal portion. In some experiments the epididymal phasic response was a single slow contraction, while in others it had a mixture of fast and slow responses. 2 Phentolamine reduced the phasic response but not the tonic response of the intact vas deferens. This effect was not observed after denervation produced by chronic guanethidine treatment. 3 Both phases of the response to KC1 160 mmol/1 were substantially reduced by phentolamine in the epididymal portion. In the prostatic portion phentolamine produced only slight inhibition of the phasic component and had no effect on the tonic component. 4 Isoprenaline had no effect on the response to KC1160 mmol/1 but reduced both phases of the response to KC150 mmol/1. This effect was antagonized by propranolol. 5 It is concluded that part of the phasic component of the response to KC1 in the rat vas deferens is due to the release of noradrenaline from intramural nerves.     

Prostaglandin-versus alpha-adrenoceptor-mediated control of sympathetic neurotransmitter secretion in guinea-pig isolated vas deferens

Noradrenaline (NA) secretion of isolated superfused guinea-pig vas deferenswas studied by determination of total field stimulation-induced efflux of tritium, after preincubation with ³H-l-NA. The medium contained optimal concentrations of desmethylimipramine and normetanephrine to block local rebinding of free NA. Further addition of the two chemically different inhibitors of prostaglandin synthetase, 5,811,14 eicosatetraynoic acid or indomethacin, consistently enhanced the nerve stimulation-induced output of tritium at a frequency of 5/sec, but not at 10/sec. The α-adrenoceptor blocking agent, phenoxybenzamine further strongly elevated nerve stimulation-induced output of tritium. This rise was abolished by low concentrations of exogenous prostaglandin E₂. The results show that sympathetic neurotransmitter secretion in the guinea-pig vas deferens, during low frequency stimulation, is restricted by local formation of prostaglandin E. However and in addition, neurotransmitter secretion appears to be restricted by an α-adrenoceptor-mediated mechanism, which does not appear to depend on endogenous prostaglandin E as a chemical mediator.     

Phenotypic and genetic characterization of patients with features of "nonclassic" forms of cystic fibrosis

OBJECTIVE: To determine which features of incomplete or "nonclassic" forms of cystic fibrosis (CF) are associated with deleterious CF transmembrane conductance regulator gene ( CFTR ) mutations, and to explore other etiologies for features not associated with deleterious CFTR mutations. STUDY DESIGN: Clinical features were compared between 57 patients with deleterious mutations in each CFTR and 63 with no deleterious mutations. The Shwachman Bodian Diamond syndrome gene ( SBDS ) was sequenced to search for mutations in patients with no deleterious CFTR mutations and steatorrhea to determine if any had unrecognized Shwachman-Diamond syndrome (SDS). RESULTS: The presence of a common CF-causing mutation, absence of the vas deferens, and Pseudomona aeruginosa in the sputum correlated with the presence of two deleterious CFTR mutations, whereas sweat chloride concentration, diagnostic criteria for CF, and steatorrhea did not. However, sweat chloride concentration correlated with CFTR mutation status in patients infected with P aeruginosa. One patient had disease-causing mutations in each SBDS . CONCLUSIONS: Presence of a common CF-causing mutation, absence of the vas deferens and/or P aeruginosa infection in a patient with features of nonclassic CF are predictive of deleterious mutations in each CFTR , whereas steatorrhea in the same context is likely to have etiologies other than CF transmembrane conductance regulator (CFTR) dysfunction.     

The effects of orchidopexy and orchidectomy on testes of rats subjected to ipsilateral abdominal testis with vas deferens obstruction

An experimental study was planned to evaluate and compare the effects of orchidopexy and orchidectomy on the testes of rats subjected to unilateral abdominal testis with vas deferens obstruction. Four groups were established. Rats in the control group underwent a sham procedure. While the testis was maintained in the abdomen with the vas deferens ligated for 8 weeks in group 2, rats in groups 3 and 4 underwent orchidopexy or orchidectomy after 4 weeks. Remaining testes were harvested at the end of the 8-week period. Testis and body weights were obtained during harvest. Samples were evaluated through DNA flow cytometry, and percentages of haploid cells were determined. Groups were compared through unpaired t-test, and p-values less than 0.05 were considered significant. All three treatments had decreased testis weight over body weight values of ipsilateral testes. Ipsilateral orchidectomy increased the value among contralateral testes. However, none of the groups had a contralateral testicular value less than the sham-operated group. All three treatments decreased the percentage of haploid cells among ipsilateral testes, but only an abdominal testis was associated with a decrease in the percentage of haploid cells among contralateral testes. Maintaining a testis with an obstructed vas deferens in the abdomen for 8 weeks damages both ipsilateral and contralateral testes. Orchidopexy, while showing amelioration of the ipsilateral testis, spares the contralateral testis as well as orchidectomy. Orchidopexy for an undescended testis with vas deferens obstruction is a rational approach.     

Ionic currents in single smooth muscle cells of the human vas deferens

PURPOSE: Smooth muscle cells of the vas deferens have an important role in carrying sperms to the exterior but little is known of their electrophysiological properties. We characterized the voltage-gated ion channel currents in single smooth muscle cells of the human vas deferens (HVSMCs). MATERIALS AND METHODS: We observed contractile responses of 8 circular smooth muscle strips of the human vas deferens to a high concentration (10 mM) of tetraethylammonium. HVSMCs were isolated using proteolytic enzymes (collagenase and papain), and were used for an electro-physiological study using whole cell and inside-out patch clamp configurations. RESULTS: The application of 10 mM tetraethylammonium induced rhythmic contractions of the strips. When HVSMCs were dialyzed with a KCl solution, step depolarizations of membrane potential evoked oscillatory outward K currents that were not inactivated. The large conductance Ca activated K (BKCa) and delayed rectifier components of the outward current were identified. The BKCa channel showed a large single channel conductance (162.7 +/- 13.2 pS with 5 mM K in the patch pipette). Two types of Ca currents were identified in the whole cell configuration. With a cell held at -50 mV an L-type Ca current was present during a depolarizing step pulse. From a holding potential of -90mV L-type and T-type Ca currents were elicited by depolarizing step pulses. CONCLUSIONS: HVSMCs have 2 (L and T) types of Ca channels and 2 types of K (BKCa and delayed rectifier) channels. Voltage dependent changes of these ion channels and their interactions may be important in regulating vas contractility.     

Relation between the anatomical genital phenotype and cystic fibrosis transmembrane conductance regulator gene mutations in the absence of the vas deferens

OBJECTIVE: To study the correlation between genital phenotype and cystic fibrosis genotype in men lacking at least one vas deferens. DESIGN: Prospective study. SETTING: Institut Rh?nalpin pour la Reproduction Humaine, Lyon-Bron, France. PATIENT(S): Forty-seven infertile men lacking at least one vas deferens. INTERVENTION(S): All patients were screened for the 13 most common CFTR gene mutations and for the 5-thymidine variant of intron 8. Renal, scrotal, and transrectal ultrasonography were systematically performed. MAIN OUTCOME MEASURE(S): Epididymal and seminal vesicular abnormalities and testicular volume were compared among men with two, one, or no CFTR gene mutation, with or without the 5T allele. RESULTS: Seminal vesicles and the symmetry of epididymal and vesicular abnormalities did not differ between patients with and those without the CFTR gene mutation. Epididymal abnormalities were more frequent in men without the mutation. Testicular volumes were significantly lower in men without the mutation and those with the 5T allele only. CONCLUSION: Men with the CFTR mutation, the 5T allele only, and those without CFTR mutation have few differences in genital phenotype. Low testicular volume is observed in men without the CFTR mutation and those with the 5T allele only.     

Alpha 2-adrenoceptor and NO mediate the opioid subsensitivity in isolated tissues of cholestatic animals

1. Our previous report showed that in acute cholestasis, the subsensitivity to morphine inhibitory effect on electrical-stimulated contractions develops significantly faster in guinea-pig ileum (GPI) and in mouse vas deferens (MVD) (45.2 and 29.9 times, respectively) compared with non-cholestatic subjects. 2. The possible contribution of alpha2-adrenoceptor and nitric oxide (NO) pathways on the development of tolerance was assessed in GPI and MVD of cholestatic subjects. 3. Daily administration of naltrexone (20 mg kg(-1)), yohimbine (5 mg kg(-1)), and Nomega-nitro-l-arginine methyl ester (l-NAME) (3 mg kg(-1)) to cholestatic animals significantly (P-value < 0.05) inhibited the process of subsensitivity in all groups. 4. Consistent with the literature, it was concluded that both the alpha2-adrenergic system and NO have close interaction with the opioid system and may underlie some of the mechanisms involved in the subsensitivity development to opioids in acute cholestatic states.