Identification and Functional Expression of Nicotinic Acetylcholine Receptors in Mcn Terminals

Chronic and acute nicotine exposure causes pregnancy and fertility problems as well as increased blood pressure and heart rate, vasoconstriction and cardiovascular disease. These physiological processes are regulated by the neuropeptides arginine vasopressin and oxytocin, which are synthesized and released from magnocellular neurons (MCNs) in the hypothalamic-neurohypophyseal system (HNS).Nicotinic acetylcholine receptors (nAChRs) are ligand gated ion channels which bind nicotine as well as the endogenous neurotransmitter acetylcholine and are widely distributed throughout the central nervous system. However, little is currently known about the distribution and function of nAChRs in the HNS. Using RT-PCR in conjunction with immunolabeling, Ca imaging and release studies, we provide a comprehensive description of the types, localization and role of nAChRs in the HNS. RT-PCR identified the presence of alpha7, alpha4, alpha2, beta4 and beta2 subunits in tissue obtained from the Supraoptic and Paraventricular nuclei; with alpha7 and beta2 being predominately expressed. Immunolabeling supports this finding and was used to quantify subunit expression and determine the anatomical location of subunits within these neurons. Immunolabeling identified alpha7, alpha4, alpha2, beta4 and beta 2 subunits in MCN terminals. The presence of alpha7 and alpha4 nAChRs was also confirmed using pharmacological and electrophysiological measurements from isolated terminals. Furthermore, neuropeptide release in the presence of nicotine and/or anatoxin was enhanced in isolated terminals and whole glands. Finally, we confirmed the presence of the vesicular acetylcholine transporter in MCN terminals indicating a possible cholinergic autocrine modulation of peptide release in the HNS. (Supported by UMass Grant P60037094900000 to SOM)