pituitrin and 5-nitro-2-(3-phenylpropylamino)benzoic-acid

Previous work on the whole neurohypophysis has shown that hypotonic conditions increase release of taurine from neurohypophysial astrocytes (pituicytes). The present work confirms that taurine is present in cultured pituicytes, and that its specific release increases in response to a hypotonic shock. We next show that vasopressin (VP) and oxytocin (OT) also specifically release taurine from pituicytes. With an EC(50) of approximately 2 nm, VP is much more potent than OT, and the effects of both hormones are blocked by SR 49059, a V(1a) receptor antagonist. This pharmacological profile matches the one for VP- and OT-evoked calcium signals in pituicytes, consistent with the fact that VP-induced taurine efflux is blocked by BAPTA-AM. However, BAPTA-AM also blocks the taurine efflux induced by a 270 mosmol l(-1) challenge, which per se does not evoke any calcium signal, suggesting a permissive role for calcium in this case. Nevertheless, the fact that structurally unrelated calcium-mobilizing agents and ionomycin are able to induce taurine efflux suggests that calcium may also play a signalling role in this event. It is widely accepted that in hypotonic conditions taurine exits cells through anionic channels. Antagonism by the chloride channel inhibitors 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) and 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) suggests the same pathway for VP-induced taurine efflux, which is also blocked in hypertonic conditions (330 mosmol l(-1)). Moreover, it is likely that the osmosensitivity of the taurine channel is up-regulated by calcium. These results, together with our in situ experiments showing stimulation of taurine release by endogenous VP, strengthen the concept of a glial control of neurohormone output.

Topics: 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Animals; Calcium; Cells, Cultured; Feedback, Physiological; Hypotonic Solutions; Intracellular Membranes; Nitrobenzoates; Pituitary Gland; Pituitary Hormones; Pituitary Hormones, Posterior; Rats; Rats, Wistar; Taurine; Vasopressins

A number of transporters and channels have been identified in cholangiocytes, but the role that bile ducts play in the formation of bile in vivo is unclear. We determined the contribution of cholangiocytes to bile flow and biliary bicarbonate excretion in normal rat liver.. Bile flow and biliary bicarbonate were measured in isolated rat livers perfused via both the portal vein and the hepatic artery because the hepatic artery provides the blood supply to bile ducts. Livers were perfused with secretin or acetylcholine (ACh), which respectively increase either adenosine 3',5'-cyclic monophosphate (cAMP) or cytosolic Ca(2+) in cholangiocytes. Livers also were perfused with glucagon or vasopressin to instead increase cAMP or cytosolic Ca(2+) in hepatocytes.. Secretin increased biliary bicarbonate in a dose-dependent fashion and was much more effective when administered via the hepatic artery. Secretin did not affect bile flow. Similarly, ACh increased bicarbonate excretion when infused via the hepatic artery but not the portal vein. The effects of secretin were augmented by ACh, and this was prevented by cyclosporin A. The effects of ACh were blocked by 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), 5-nitro2-(3-phenylpropylamino)benzoic acid (NPPB), or diphenylamine-2-carboxylic acid (DPC), and the effects of secretin were inhibited by NPPB or DPC and unaffected by DIDS. Neither glucagon nor vasopressin altered biliary bicarbonate.. Biliary bicarbonate is regulated by cholangiocytes rather than hepatocytes in normal rat liver. ACh-induced bicarbonate excretion depends on both chloride channels and bicarbonate exchange, whereas secretin-induced bicarbonate excretion is independent of bicarbonate exchange.

Topics: 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Acetylcholine; Angiogenesis Inhibitors; Animals; Anti-Inflammatory Agents, Non-Steroidal; Bicarbonates; Bile; Bile Ducts; Biological Transport; Calcium; Cyclic AMP; Cyclosporine; Dose-Response Relationship, Drug; Enzyme Inhibitors; Epithelial Cells; Glucagon; Hepatic Artery; Hepatic Veins; In Vitro Techniques; Liver; Male; Nitrobenzoates; ortho-Aminobenzoates; Rats; Rats, Sprague-Dawley; Secretin; Vasoconstrictor Agents; Vasodilator Agents; Vasopressins

Osmotic regulation of supraoptic nucleus (SON) neuron activity depends in part on activation of neuronal glycine receptors (GlyRs), most probably by taurine released from adjacent astrocytes. In the neurohypophysis in which the axons of SON neurons terminate, taurine is also concentrated in and osmo-dependently released by pituicytes, the specialized glial cells ensheathing nerve terminals. We now show that taurine release from isolated neurohypophyses is enhanced by hypo-osmotic and decreased by hyper-osmotic stimulation. The high osmosensitivity is shown by the significant increase on only 3.3% reduction in osmolarity. Inhibition of taurine release by 5-nitro-2-(3-phenylpropylamino)benzoic acid, niflumic acid, and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid suggests the involvement of volume-sensitive anion channels. On purified neurohypophysial nerve endings, activation of strychnine-sensitive GlyRs by taurine or glycine primarily inhibits the high K(+)-induced rise in [Ca(2+)](i) and subsequent release of vasopressin. Expression of GlyRs in vasopressin and oxytocin terminals is confirmed by immunohistochemistry. Their implication in the osmoregulation of neurohormone secretion was assessed on isolated whole neurohypophyses. A 6.6% hypo-osmotic stimulus reduces by half the depolarization-evoked vasopressin secretion, an inhibition totally prevented by strychnine. Most importantly, depletion of taurine by a taurine transport inhibitor also abolishes the osmo-dependent inhibition of vasopressin release. Therefore, in the neurohypophysis, an osmoregulatory system involving pituicytes, taurine, and GlyRs is operating to control Ca(2+) influx in and neurohormone release from nerve terminals. This elucidates the functional role of glial taurine in the neurohypophysis, reveals the expression of GlyRs on axon terminals, and further defines the role of glial cells in the regulation of neuroendocrine function.

Topics: 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Animals; Calcium; Chloride Channels; Glycine; Glycine Agents; Immunohistochemistry; In Vitro Techniques; Male; Neuroglia; Niflumic Acid; Nitrobenzoates; Osmolar Concentration; Oxytocin; Pituitary Gland, Posterior; Presynaptic Terminals; Rats; Rats, Wistar; Receptors, Glycine; Supraoptic Nucleus; Taurine; Vasopressins

The effects of aldosterone and vasopressin on Cl- transport were investigated in a mouse cortical collecting duct (mpkCCD) cell line derived from a transgenic mouse carrying the SV40 large T antigen driven by the proximal regulatory sequences of the L-pyruvate kinase gene. The cells had features of a tight epithelium and expressed the amiloride-sensitive sodium channel and the cystic fibrosis transmembrane conductance regulator (CFTR) genes. dD-arginine vasopressin (dDAVP) caused a rapid, dose-dependent, increase in short-circuit current (Isc). Experiments with ion channel blockers and apical ion substitution showed that the current represented amiloride-sensitive Na+ and 5-nitro-2-(3-phenylpropylamino)benzoate-sensitive and glibenclamide-sensitive Cl- fluxes. Aldosterone (5 x 10(-7)M for 3 or 24 hr) stimulated Isc and apical-to-basal 22Na+ flux by 3-fold. 36Cl- flux studies showed that dDAVP and aldosterone stimulated net Cl- reabsorption and that dDAVP potentiated the action of aldosterone on Cl- transport. Whereas aldosterone affected only the apical-to-basal 36Cl- flux, dDAVP mainly increased the apical-to-basal Cl- flux and the basal-to-apical flux of Cl- to a lesser extent. These results suggest that the discrete dDAVP-elicited Cl- secretion involves the CFTR and that dDAVP and aldosterone may affect in different ways the observed increased Cl- reabsorption in this model of mouse cultured cortical collecting duct cells.

Topics: Aldosterone; Amiloride; Animals; Cell Line, Transformed; Chloride Channels; Cystic Fibrosis Transmembrane Conductance Regulator; Deamino Arginine Vasopressin; Epithelial Sodium Channels; Gene Expression; Kidney Tubules, Collecting; Male; Mice; Mice, Transgenic; Nitrobenzoates; Sodium Channels; Sodium Radioisotopes; Vasopressins

Agonists which stimulate the inositol 1,4,5 trisphosphate ([1,4,5]-IP3)-dependent mobilization of Ca2+ from intracellular stores also stimulate entry of divalent cations across the cell membrane. Under appropriate experimental conditions, divalent cation entry across the cell membrane can be monitored as the rate at which the intracellular fluorescence of divalent cation indicators is quenched by the addition of Mn2+ to the extracellular medium. We report that addition of vasopressin to fura-2-loaded glomerular mesangial cells in culture markedly accelerated the rate at which Mn2+ quenched fura-2 fluorescence at its Ca(2+)-insensitive wavelength in the presence of extracellular NaCl, but that this quench response was attenuated when Cl- was removed from the extracellular medium by equimolar substitution with impermeant anions (gluconate, methanesulfonate, acetate, lactate). Similarly, loss of agonist-induced quench also occurred when Cl- was substituted with gluconate in K(+)-containing media. Addition of the Cl- channel inhibitor, 5-nitro-2-(3-phenylpropylaminobenzoic acid) (NPPB), also inhibited Mn(2+)-induced quench of fura-2 fluorescence following vasopressin addition. In contrast, in the presence of gramicidin to provide an alternate conductance pathway to accompany divalent cation entry, agonist-dependent Mn2+ quench occurred even in the absence of extracellular Cl-, indicating that the requirement for Cl- was not the result of cotransport on a common transporter nor the result of Cl- serving as a necessary cofactor for divalent cation entry. A similar dependence on extracellular Cl- was observed for other Ca(2+)-mobilizing agonists such as endothelin, as well as the intracellular Ca2+ ATPase inhibitor, thapsigargin. Extracellular Cl- dependence for agonist-induced divalent cation entry was also reflected in a corresponding extracellular Cl- dependence for agonist-induced mesangial cell contraction. It has been previously shown by ourselves (Kremer et al., 1992a, Am. J. Physiol., 262:F668-F678) and others that agonist-stimulated calcium mobilization in mesangial cells is accompanied by inhibition of K+ conductance and increased Cl- conductance. Accordingly, we conclude that the current findings suggest that activation of Cl- conductance provides regulated charge compensation for receptor-mediated divalent cation entry in response to Ca(2+)-mobilizing vasoconstrictor agonists in mesangial cells.

Topics: Animals; Calcium-Transporting ATPases; Cations, Divalent; Cell Membrane Permeability; Cells, Cultured; Chloride Channels; Chlorides; Endothelins; Fluorescence; Glomerular Mesangium; Gluconates; Gramicidin; Hydrogen-Ion Concentration; Manganese; Nitrobenzoates; Potassium Channels; Rats; Rats, Sprague-Dawley; Receptors, Cell Surface; Sodium Chloride; Terpenes; Thapsigargin; Vasopressins

We have investigated the cellular signalling pathway by which vasopressin stimulates a Ca2(+)-dependent Cl- conductance and the effects of two known Cl- channel blockers in cultured rat A7r5 aortic smooth muscle cells using anion efflux and fluorescent Ca2+ imaging studies. Addition of vasopressin (100 nM) to A7r5 cells enhanced 125I (Cl- substitute) efflux from the cells through a V1 receptor-mediated pathway. Maximal increases in the rate of efflux were observed 1 min following addition of vasopressin (4-fold above basal levels). Activation of the V1 pathway was demonstrated by an increase in inositol trisphosphate (IP3) formation and lack of cAMP accumulation by the cells following the addition of vasopressin. Fluorescent ratio imaging with fura-2 revealed that addition of vasopressin to the cells results in an increase of [Ca2+]i which peaks within 20 s and does not return to resting levels during the 100 s observation period. The addition of a Ca2+ ionophore mimicked the vasopressin-induced efflux from the cells. 5-Nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) and a chloro-substituted compound (cpd 149) inhibited the vasopressin-stimulated 125I efflux from the cells. The concentrations of NPPB and cpd 149 required to inhibit 125I efflux from the cells were similar to those which also attenuated vasopressin-induced Ca2+ transients in the cells. NPPB and cpd 149 had no effects on the ionomycin stimulated efflux. The mechanism(s) by which cpd 149 exerts its effect on stimulated efflux was examined by measuring its action on vasopressin-induced changes in IP3. Compound 149 inhibited IP3 generation in response to vasopressin.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Aorta, Thoracic; Calcium; Cell Line; Chloride Channels; Colforsin; Cyclic AMP; Homeostasis; Inositol Phosphates; Iodine; Iodine Radioisotopes; Membrane Proteins; Muscle, Smooth, Vascular; Nitrobenzoates; ortho-Aminobenzoates; Rats; Vasopressins