Orexins/Hypocretins Acting at G sub(i) Protein-Coupled OX sub(2) Receptors Inhibit Cyclic AMP Synthesis in the Primary Neuronal Cultures

• Published in: Journal of molecular neuroscience Vol. 46; no. 1; pp. 10 - 17
• Main Authors: Urbaska, Anna, Sokoowska, Paulina, Woldan-Tambor, Agata, Biegaska, Kaja, Brix, Britta, Johren, Olaf, Namieciska, Magdalena, Zawilska, Jolanta Barbara
• Format: Journal Article
• Language: English
• Published: 01.01.2012
• Subjects: Adenylate cyclase; Cell culture; Cortex; Cyclic AMP; Forskolin; G protein-coupled receptors; Nervous system; Neuropeptides; Nucleotides; Orexin receptors; Orexins; pertussis toxin; Pituitary adenylate cyclase-activating polypeptide; Receptor mechanisms; Vasoactive intestinal peptide
• ISSN: 0895-8696; 1559-1166
• DOI: 10.1007/s12031-011-9526-2

Orexins A and B are newly discovered neuropeptides with pleiotropic activity. They signal through two G protein-coupled receptors: OX sub(1) and OX sub(2). In this study, we examined the expression of orexin receptors and effects of the receptors' activation on cyclic AMP formation in the primary neuronal cell cultures from rat cerebral cortex. Both types of orexin receptors were expressed in rat cortical neurons; the level of OX sub(2)R was markedly higher compared to OX sub(1)R. Orexin A (an agonist of OX sub(1)R and OX sub(2)R) and [Ala super(11)-D-Leu super(15)]orexin B (a selective agonist of OX sub(2)R) did not affect basal cyclic AMP formation in the primary neuronal cell cultures. Both peptides (0.001-1 mu M) inhibited, in a concentration-dependent manner and IC sub(50) values in low nanomolar range, the increase in the nucleotide production evoked by forskolin (1 mu M; a direct activator of adenylyl cyclase), pituitary adenylate cyclase-activating polypeptide (PACAP27; 0.1 mu M), and vasoactive intestinal peptide (VIP; 3 mu M). Effects of orexin A on forskolin-, PACAP27-, and VIP-stimulated cyclic AMP synthesis were blocked by TCS OX2 29 (a selective antagonist of OX sub(2)R), and unaffected by SB 408124 (a selective antagonist of OX sub(1)R). Pretreatment of neuronal cell cultures with pertussis toxin (PTX) abolished the inhibitory action of orexin A on forskolin- and PACAP-stimulated cyclic AMP accumulation. It is suggested that in cultured rat cortical neurons orexins, acting at OX sub(2) receptors coupled to PTX-sensitive G sub(i) protein, inhibit cyclic AMP synthesis.