Activation of cystic fibrosis transmembrane conductance regulator in rat epididymal epithelium by genistein

The effect of genistein on anion secretion via cystic fibrosis transmembrane conductance regulator (CFTR) in cultured rat cauda epididymal epithelia was studied by short-circuit current (Isc) technique. Genistein added apically stimulated a concentration-dependent rise in Isc due to Cl- and HCO3 - secretion. The genistein-induced Isc was observed in basolaterally permeabilized monolayers, suggesting that the Isc response was mediated by the apical anion channel. The response could be blocked by the nonspecific Cl- channel blocker, diphenylamine-2-carboxylate (DPC), but not by the Ca2+-activated Cl- channel blocker, 4,4'-diisothiocyanostilbene- 2,2'-disulfonic acid (DIDS). Genistein did not increase intracellular cAMP, but H-89, a protein kinase A inhibitor, completely abolished the Isc response to genistein. Moreover, pretreatment of the tissues with MDL-12330A, an adenylate cyclase inhibitor, markedly attenuated the Isc response to genistein, but the response was restored upon the addition of exogenous cAMP. Ca2+, protein kinase C, tyrosine kinase, and protein phosphatase signalling pathways were not involved in the action of genistein. It is speculated that genistein stimulates anion secretion by direct interaction with CFTR. This requires a low level of phosphorylation of CFTR by basal protein kinase A activity. It is suggested that genistein may provide therapeutic benefit to male infertility associated with cystic fibrosis.link_to_subscribed_fulltex

A method for studying apical anion conductance in cultured rat epididymal cells using the short-circuit current technique

79P–138P of this journal issue entitled: Proceedings of the Physiological Society, 18-20 July 1991, Cambridge Meeting: Communications: Part 1link_to_OA_fulltex

Immunoactive TSH in the amniotic fluid of the rat

Amniotic fluid was obtained from 19-day-old rat fetuses by aspiration. Pooled samples measured at 4 different dilutions demonstrated parallelism with standard rat TSH. It is concluded that rat amniotic fluid has TSH immunoactivity.link_to_subscribed_fulltex

Ultrastructural correlation of water reabsorption in isolated rat cauda epididymidis

Electron microscopic study was made on the water reabsorption of the epithelial cells of the rat cauda epididymidis. It was shown that when the epididymal duct was reabsorbing water at a maximal rate, widely dilated intercellular spaces were seen. It is suggested that the standing gradient model of water reabsorption first proposed for the gall bladder may also operate in the cauda epididymidis.link_to_subscribed_fulltex

Serum potassium and aldosterone levels in gossypol-treated rats

Male rats fed with 30 mg/kg gossypol for 2 or 4 weeks exhibited a decrease in growth but showed no change from control in serum potassium levels. Serum aldosterone levels remained unchanged, but serum testosterone levels were significantly reduced after 4 weeks of treatment. Serum T3 was significantly increased after or 4 weeks of treatment, while serum T4 was not different from control. It is concluded that gossypol at a dose that is sufficient to produce sterility in rats did not change serum potassium and aldosterone levels.link_to_subscribed_fulltex

Biphasic short-circuit current response to noradrenalin mediated by Ca 2+ and cAMP in cultured rat epididymal epithelium

A study was carried out to investigate the short-circuit current (I(SC)) response to noradrenaline (NA) and the signal transduction mechanisms involved in cultured rat cauda epididymal epithelium. In normal Krebs-Henseleit solution, NA (10 μmol·l -1) added basolaterally elicited a biphasic I(SC) response consisting of a transient spike followed by a second sustained response. The biphasic response was almost abolished by removing ambient Cl -. Preloading the tissues with a cell permeant Ca 2+ chelator, 1,2-bis(2-aminophenoxy) ethane-N,N,N',N',-tetraacetic acid acetoxymethyl ester (BAPTA/AM), or pretreating them with thapsigargin (Tg), a microsomal adenosine triphosphatase inhibitor abolished the initial spike in the I(SC) response to NA, but had little effect on the second component. Pretreating the tissues with a non-selective β-antagonist, nadolol, reduced the second I(SC) response in a dose-dependent fashion but the initial spike was not affected. Microfluorimetric studies showed that NA (100 μmol·l -1) elicited single Ca 2+ spikes in isolated epididymal cells, which could be abolished by prior treatment with Tg. Biochemical assays showed that NA (10 μmol·l -1) increased intracellular cyclic adenosine monophosphate concentration ([cAMP](i)) and the response was abolished by prior treatment with nadolol (50 μmol·l -1). The results showed that NA elicited a biphasic I(SC) response mediated by a rise in intracellular Ca 2+ concentration ([Ca 2+](i)) followed by a rise in [cAMP](i). The Ca 2+-mediated I(SC) response had a faster onset and more transient action than the cAMP counterpart. It is suggested that NA released from noradrenergic nerve endings regulates transepithelial Cl - secretion in the epididymis thereby providing the specialized milieu vital for sperm storage and maturation.link_to_subscribed_fulltex

Ca 2+ release in cultured rat epididymal cells during hypoosmotic swelling

Microfluorimetric studies were carried out to investigate the effects of hypoosmotic swelling on intracellular Ca 2+ concentration ([Ca 2+](i)) in single rat epididymal cells. In Ca 2+-free solution containing 50 μmol/l ethylenebis(oxonitrilo)tetraacetate (EGTA) hypoosmotic swelling (-160 mosmol/l) induced a transient rise in [Ca 2+](i)< which was either monophasic, biphasic or oscillatory. The [Ca 2+](i) responses to repeated hypoosmotic stimulations followed a decremental pattern. However, if 2.5 mmol/l Ca 2+ was admitted during the recovery period between successive stimulations, the second and the third [Ca 2+](i) responses were slightly greater than the first. Increasing the change in osmolarity from -14 ± 1.0 to -154 ± 1.5 mosmol/l increased the rise in [Ca 2+](i) but reduced the [Ca 2+](i) response to subsequent ionomycin stimulation (4 μmol/l). The swelling- and the ionomycin-induced rises in [Ca 2+](i) followed a reciprocal pattern. It was suggested that intracellular Ca 2+ release in response to cell swelling in the epididymal epithelium might play a role in cell volume regulation and the control of epididymal fluid osmolarity.link_to_subscribed_fulltex

Nongenomic effect of testosterone on chloride secretion in cultured rat efferent duct epithelia

Short-circuit current (Isc) technique was used to investigate the role of testosterone in the regulation of chloride secretion in cultured rat efferent duct epithelia. Among the steroids tested, only testosterone, and to a lesser extent, 5α-dihydrotestosterone (5α-DHT), reduced the basal and forskolin-induced Isc in cultured rat efferent duct epithelia when added to the apical bathing solution. Indomethacin, a 3α-hydroxysteroid dehydrogenase, did not affect the inhibitory effect of 5α-DHT. The effect of testosterone occurred within 10-20 s upon application and was dose dependent with apparent IC50 value of 1 μM. The effect was abolished by removal of C1- but not HCO3 - from the normal Krebs-Henseleit solution, suggesting that testosterone mainly inhibited C1- secretion. The efferent duct was found to be most sensitive to testosterone, while the caput and the cauda epididymidis were only mildly sensitive. Cyproterone acetate, a steroidal antiandrogen, or flutamide, a nonsteroidal antiandrogen, did not block the effect of testosterone on the forskolin-induced Isc, nor did protein synthesis inhibitors, cycloheximide, or actinomycin D. However, pertussis toxin, a Gi protein inhibitor, attenuated the inhibition of forskolin-induced Isc by testosterone. Testosterone caused a dose-dependent inhibition of forskolin-induced rise in cAMP in efferent duct cells. It is suggested that the rapid effect of testosterone was mediated through a membrane receptor that is negatively coupled to adenylate cyclase via Gi protein. The role of nongenomic action of testosterone in the regulation of electrolyte and fluid transport in the efferent duct is discussed.link_to_subscribed_fulltex