1. [Arg8]-vasopressin-induced increase in intracellular Ca2+ concentration in cultured rat hippocampal neurons
T Mihara, T Tarumi, Y Sugimoto, Z Chen, C Kamei Brain Res Bull. 1999 Jul 15;49(5):343-7. doi: 10.1016/s0361-9230(99)00064-7.
Changes in intracellular Ca2+ concentration ([Ca2+]i) induced by [Arg8]-vasopressin (AVP) were studied in cultured rat hippocampal neurons by fura-2 fluorometry. AVP (10-1,000 nM) caused a dose-dependent increase in [Ca2+]i. The selective V1 vasopressin receptor agonist [Phe2, Ile3, Orn8]-vasopressin also induced a significant increase in [Ca2+]i, whereas the selective V2 vasopressin receptor agonist [deamino Cys1, D-Arg8]-vasopressin showed no effect. The AVP-induced increase in [Ca2+]i was inhibited by the selective V1 vasopressin receptor antagonist d(CH2)5[Tyr2(Me), Arg8]-vasopressin and nonpeptide V1 antagonist OPC-21268. On the other hand, no antagonistic effects were observed with the V2 vasopressin antagonist desglycinamide-[d(CH2)5, D-Ile2, Ile4, Arg8]-vasopressin and nonpeptide V2 antagonist OPC-31260. The increase in [Ca2+]i induced by AVP was abolished after removal of extracellular Ca2+. In addition, AVP-induced [Ca2+]i elevation was not affected by treatment with verapamil, which blocked the [Ca2+]i increase induced by an isotonic high K(+)-medium (50 mM). However, omega-conotoxin GVIA completely inhibited the effect of AVP. These results suggested that the AVP-induced [Ca2+]i increase in cultured rat hippocampal neurons is due to influx of Ca2+ through V1 VP receptors coupled with N-type calcium channels.
2. Neurohypophyseal peptides and tachykinins stimulate the production of cyclic GMP in cultured porcine aortic endothelial cells
V B Schini, Z S Katusic, P M Vanhoutte J Pharmacol Exp Ther. 1990 Dec;255(3):994-1000.
The response to small peptides such as Arg-vasopressin, oxytocin and tachykinins was investigated in cultured porcine aortic endothelial cells. The production of endothelium-derived nitric oxide was assessed indirectly by the accumulation of cyclic GMP, a response that is due to the increased activity of soluble guanylate cyclase of the endothelial cells after release of the mediator. Arg-vasopressin, oxytocin, substance P and physalae-min (an analog of substance P, pGlu-Ala-Asp-Pro-Asn-Lys-Phe-Tyr-Gly-Leu-Met-NH2) markedly and transiently stimulated the production of cyclic GMP without affecting that of cyclic AMP. Treatment of endothelial cells with either hemoglobin or methylene blue reduced significantly both the basal and stimulated level of cyclic GMP. The production of cyclic GMP evoked by Arg-vasopressin and substance P was inhibited selectively by NG-monomethyl-L-arginine but not by its D-enantiomer. The neurohypophyseal hormones and related peptides stimulated the accumulation of cyclic GMP in a concentration-dependent manner, with the following relative order of potency: oxytocin greater than Lys-vasopressin greater than Arg-vasopressin much greater than [deamino-Cys1, D-Arg8]-vasopressin. The production of cyclic GMP evoked by oxytocin was inhibited selectively by [d(CH2)5, Tyr(OMe)2, Orn8]-vasotocin, an oxytocin antagonist. The production of cyclic GMP evoked by Arg-vasopressin and Lys-vasopressin was inhibited by [beta-mercapto-beta, beta-cyclopentamethylene-propionyl1, O-Me-Tyr2, Arg8]-vasopressin, a selective V1-receptor antagonist. The moderate production of cyclic GMP evoked by [deamino-Cys1, D-Arg8]-vasopressin was inhibited significantly by the V1-receptor antagonist. The peptide antagonists affected only minimally or not at all the production of cyclic GMP evoked by a donor of nitric oxide, SIN-1 (3-Morpholino-Sydnonimine). These observations indicate that 1) neurohypophyseal hormones and tachykinins stimulate the accumulation of cyclic GMP in cultured porcine aortic endothelial cells by increasing the production of endothelial-derived nitric oxide, which in turn enhances the activity of soluble guanylate cyclase; 2) the production of cyclic GMP in response to oxytocin is due to activation of oxytocinergic receptors; and 3) the production of cyclic GMP evoked by Arg-vasopressin and Lys-vasopressin is due mostly to activation of V1-vasopressinergic receptors.
3. Muscle layer- and region-dependent distributions of oxytocin receptors in the porcine myometrium
T Kitazawa, T Kajiwara, A Kiuchi, H Hatakeyama, T Taneike Peptides. 2001 Jun;22(6):963-74. doi: 10.1016/s0196-9781(01)00402-8.
The aim of the present study was to clarify smooth muscle- and region-dependent distributions of the oxytocin receptor that mediates oxytocin-induced contraction in the nonpregnant porcine myometrium by means of mechanical and radioligand ([3H]-oxytocin) binding studies. In Krebs solution, oxytocin (0.1-300 nM) caused concentration-dependent contractions of the cornual myometrium, and the longitudinal muscle was more sensitive than the circular muscle. [Arg8]-vasopressin and [deamino-Cys1, D-Arg8]-vasopressin also contracted the myometrium, and the order of the potency was oxytocin > [Arg8]-vasopressin > [deamino-Cys(1), D-Arg(8)]-vasopressin. Treatment with a high concentration of oxytocin selectively inhibited the contraction of oxytocin and [Arg8]-vasopressin without affecting the responses of acetylcholine and high-K+. Selective cross inhibition was also observed in the presence of a high concentration of [Arg(8)]-vasopressin. The oxytocin-induced contraction was resistant to tetrodotoxin and atropine, but was reduced by verapamil or by the removal of external Ca2+, indicating that oxytocin has a direct action on smooth muscle cells and that extracellular Ca2+ plays an important role for the contraction. In Kumagai solution, oxytocin caused contraction of the cornual longitudinal muscle (-logEC50 = 8.5) but not the circular muscle. Longitudinal muscles of other regions (corpus and cervix) were also responsive to oxytocin, but the -logEC50 value differed from region to region (cornua > corpus = cervix). On the other hand, oxytocin failed to cause contraction of the corpus and cervical circular muscles. 3H-Oxytocin bound to crude membrane preparations of the myometrium in a concentration-dependent (0.084-2.7 nM) saturable manner. Scatchard analysis of equilibrium binding data revealed the presence of a single class of binding site with an apparent dissociation constant (Kd, 1.1-1.5 nM), but receptor density (Bmax) differed in the two muscle layer types (longitudinal muscle: circular muscle = 5:1) and tended to decrease from the cornua to the cervix. In conclusion, the receptor specific for oxytocin is present in the porcine myometrium and mediates the contractile responses of both oxytocin and [Arg8]-vasopressin. The distribution of the oxytocin receptors differs according to the type of muscle layer (longitudinal muscle > circular muscle) and the region of the uterus.
