Dopamine D2 receptors signal through the pertussis toxin (PTX)-sensitive G(i/o) and PTX-insensitive G(z) proteins, as well as through a G protein-independent, beta-arrestin/glycogen synthase kinase-3-dependent pathway. Activation of these receptors in pituitary lactotrophs leads to inhibition of prolactin (PRL) release. It has been suggested that this inhibition occurs through the G(i/o)-alpha protein-mediated inhibition of cAMP production and/or G(i/o)-betagamma dimer-mediated activation of inward rectifier K(+) channels and inhibition of voltage-gated Ca(2+) channels. Here we show that the dopamine agonist-induced inhibition of spontaneous Ca(2+) influx and release of prestored PRL was preserved when cAMP levels were elevated by forskolin treatment. We further observed that dopamine agonists inhibited both spontaneous and depolarization-induced Ca(2+) influx in untreated but not in PTX-treated cells. This inhibition was also observed in cells with blocked inward rectifier K(+) channels, suggesting that the dopamine effect on voltage-gated Ca(2+) channel gating is sufficient to inhibit spontaneous Ca(2+) influx. However, agonist-induced inhibition of PRL release was only partially relieved in PTX-treated cells, indicating that dopamine receptors also inhibit exocytosis downstream of voltage-gated Ca(2+) influx. The PTX-insensitive step in agonist-induced inhibition of PRL release was not affected by the addition of wortmannin, an inhibitor of phosphatidylinositol 3-kinase, and lithium, an inhibitor of glycogen synthase kinase-3, but was attenuated in the presence of phorbol 12-myristate 13-acetate, which inhibits G(z) signaling pathway in a protein kinase C-dependent manner. Thus, dopamine inhibits basal PRL release by blocking voltage-gated Ca(2+) influx through the PTX-sensitive signaling pathway and by desensitizing Ca(2+) secretion coupling through the PTX-insensitive and protein kinase C-sensitive signaling pathway.

Anterior pituitary cells fire action potentials and release cyclic nucleotides both spontaneously and in response to agonist stimulation, but the relationship between electrical activity and cyclic nucleotide efflux has not been studied. In these cells, a tetrodotoxin-resistant background Na ϩ conductance is critical for firing of action potentials, and multidrug resistance proteins (MRPs) MRP4 and MRP5 contribute to cyclic nucleotide efflux. Here, we show that abolition of the background Na ϩ conductance in rat pituitary cells by complete or partial replacement of extracellular Na ϩ with organic cations or sucrose induced a rapid and reversible hyperpolarization of cell membranes and inhibition of action potential firing, accompanied by a rapid inhibition of cyclic nucleotide efflux. Valinomycin-induced hyperpolarization of plasma membranes also inhibited cyclic nucleotide efflux, whereas depolarization of cell membranes induced by the inhibition of Ca 2ϩ influx or stimulation of Na ϩ influx by gramicidin was accompanied by a facilitation of cyclic nucleotide efflux. In contrast, inhibition of cyclic nucleotide efflux by probenecid did not affect the background Na ϩ conductance. In human embryonic kidney 293 cells stably transfected with human MRP4 or MRP5, replacement of bath Na ϩ with organic cations also hyperpolarized the cell membranes and inhibited cyclic nucleotide efflux. In these cells, the Na

Gi/o protein-coupled receptors, signaling through G protein-dependent and -independent pathways, have prominent effects on secretion by modulating calcium signaling and regulating the size of the releasable secretory pool, the rates of exocytosis and endocytosis, and de novo synthesis. Pituitary cells fire action potentials spontaneously, and the associated calcium influx is sufficient to maintain prolactin (PRL) release, but not gonadotropin release, at high and steady levels for many hours. Such secretion, termed intrinsic, spontaneous or basal, reflects fusion of secretory vesicles triggered by the cell type-specific pattern of action potentials. In lactotrophs, activation of endothelin ETA and dopamine D2 receptors causes inhibition of spontaneous electrical activity and basal adenylyl cyclase activity, accompanied with inhibition of basal PRL release. Agonist-induced inhibition of cAMP production and firing of action potentials is abolished in cells with blocked pertussis toxin (PTX)-sensitive Gi/o signaling pathway. However, agonist-induced inhibition of PRL release is only partially relieved in such treated cells, indicating that both receptors also inhibit exocytosis downstream of cAMP/calcium signaling. The PTX-insensitive step in agonist-induced inhibition of PRL release is not affected by inhibition of PI3-kinase and GSK-3, but is partially rescued by down-regulation of the Gzα expression. Thus, ETA and D2 receptors inhibit basal PRL release not only by blocking electrical activity but also by desensitizing calcium-secretion coupling.

Abstract. Stress interferes with reproduction, adversely influencing implantation and fetal growth, and sometimes even leading to abortion. Here, we attempted to evaluate the early gestational effects of uncomfortable sound on pregnant mice and their offspring. Ten-week-old pregnant Jcl:ICR mice were exposed to sound (100 dB, random frequency between 9-34 kHz) for 8 hours on the 3 rd , 5 th and 7 th gestational days (GD). The effects of general anesthesia were also investigated, with or without acoustic stress. All groups were examined on the 18 th GD for fetal growth. Fetal weight, number of ossified sacrococcygeal vertebrae and placental weight were all significantly reduced (P<0.0001) when stress was induced on the 7 th GD, but not on the 3 rd or 5 th GD. This intra-uterine growth retardation (IUGR) was significantly inhibited by general anesthesia (P<0.0001), although general anesthesia alone induced significant IUGR (P<0.0001) when compared with control mice. This suggests that acoustic exposure indirectly exerts an effect on fetal growth, possibly via a psycho-maternal pathway. We also found that analysis of the number of ossified sacrococcygeal vertebrae is the most sensitive tool for the study of IUGR.

ABSTRACT-Water extract from commercial English tea has a potent inhibitory activity against human placenta aldose reductase (NADPH oxidoreductase, E.C.1.1.1.21.). Inhibitory activity was separated into five major fractions by one-step chromatography with a C-18 reverse phase column. The most active fraction was further subjected to reverse phase column chromatography. As a result, a well-known flavone-glycoside, isoquercitrin, was isolated as the most potent chemical. The inhibitory character of isoquercitrin for aldose reductase was a mix of uncompetitive and noncompetitive inhibitions, and its IC50 was 1´10 -6 M. In rat sciatic nerve tissue preparations, sorbitol accumulation in the presence of high concentrations of glucose (30 mM) was inhibited by 38% at 5´10 -4 M of isoquercitrin. The flavone-glycoside isoquercitrin is the active inhibitor of aldose reductase inhibitor present in English tea. Given the ability of aldose reductase inhibitors to prevent diabetic complications, an epidemiological study of the effect of tea consumption on the pathogenesis and progression of diabetic complications would be interesting.

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