title: Vasopressin-induced calcium signaling in cultured cortical neurons.
abstract: Earlier autoradiographic studies from our laboratory detected vasopressin recognition sites in the mammalian cerebral cortex [R.E. Brinton, K.W. Gee, J.K. Wamsley, T.P. Davis, H.I. Yamamura, Regional distribution of putative vasopressin receptors in rat brain and pituitary by quantitative autoradiography, Proc. Natl. Acad. Sci. U. S.A., 81 (1984) 7248-7252; C. Chen, R.D. Brinton, T.J. Shors, R.F. Thompson, Vasopressin induction of long-lasting potentiation of synaptic transmission in the dentate gyrus, Hippocampus, 3 (1993) 193-204]. More recently, we have detected mRNA for the V1a vasopressin receptors (V1aRs) in cultured cortical neurons [R.S. Yamazaki, Q. Chen, S.S. Schreiber, R.D. Brinton, V1a Vasopressin receptor mRNA expression in cultured neurons, astroglia, and oligodendroglia of rat cerebral cortex, Mol. Brain Res., 45 (1996) 138-140]. To determine whether these recognition sites are functional receptors, we have pursued the signal transduction mechanism associated with the V1a vasopressin receptor in enriched cultures of cortical neurons. Results of these studies demonstrate that exposure of cortical neurons to the selective V1 vasopressin receptor agonist, [Phe2,Orn8]-vasotocin, (V1 agonist) induced a significant accumulation of [3H]inositol-1-phosphate ([3H]IP1). V1 agonist-induced accumulation of [3H]IP1 was concentration dependent and exhibited a linear dose response curve. Time course analysis of V1 agonist-induced accumulation of [3H]IP1 revealed a significant increase by 20 min which then decreased gradually over the remaining 60 min observation period. V1 agonist-induced accumulation of [3H]IP1 was blocked by a selective V1a vasopressin receptor antagonist, (Phenylac1, D-Tyr(Me)2, Arg6,8, Lys-NH29)-vasopressin. Results of calcium fluorometry studies indicated that V1 agonist exposure induced a marked and sustained rise in intracellular calcium which was abolished in the absence of extracellular calcium. The loss of the rise in intracellular calcium was not due to a failure to induce PIP2 hydrolysis since activation of the phosphatidylinositol pathway occurred in the absence of extracellular calcium. V1 agonist activation of calcium influx was then investigated. V1 agonist-induced 45Ca2+ uptake was concentration dependent with a biphasic time course at 250 nM. Preincubation with the L-type calcium channel blocker, nifedipine, blocked V1 agonist-induced calcium influx suggesting V1 agonist-induced L-type calcium channel activation in cortical neurons. Furthermore, V1 agonist-induced calcium influx was blocked by both bisindolyleimide I (PKC inhibitor) and U-73122 (PLC inhibitor) suggesting a modulation of V1 agonist-induced L-type calcium channel activation by downstream components of the phosphatidylinositol signaling pathway such as protein kinase C. These results indicate that in cultured cortical neurons, V1a vasopressin receptor activation leads to induction of the phosphatidylinositol signaling pathway, influx of extracellular calcium via L-type calcium channel activation, and a rise in intracellular calcium which is dependent on V1a receptor activated influx of extracellular calcium. These data are the first to demonstrate an effector mechanism for the V1 vasopressin receptor in the cerebral cortex and provide a potential biochemical mechanism that may underlie vasopressin enhancement of memory function.
authors.affiliation: Department of Molecular Pharmacology and Toxicology, USC, 1985 Zonal Ave., Los Angeles, CA 90033, USA.
authors.forename: M C
authors.initials: MC
authors.lastname: Son
authors.forename: R D
authors.initials: RD
authors.lastname: Brinton
mesh_terms: D000818:Animals; D065092:Antidiuretic Hormone Receptor Antagonists; D001127:Arginine Vasopressin / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects* / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D005333:Fetus; D007424:Intracellular Fluid / Q000378:metabolism; D009474:Neurons / Q000502:physiology*; D010716:Phosphatidylinositols / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017483:Receptors, Vasopressin / Q000819:agonists / Q000378:metabolism
chemical_list: D065092:Antidiuretic Hormone Receptor Antagonists; D015220:Calcium Channels; D010716:Phosphatidylinositols; D017483:Receptors, Vasopressin; D001127:Arginine Vasopressin; D002118:Calcium