Immunoreactive localisation of P2Y 1 receptors within the rat and human nodose ganglia and rat brainstem: comparison with [α 33P]deoxyadenosine 5′-triphosphate autoradiography

Abstract

The present study employed standard peroxidase immunohistochemistry to map the distribution of P2Y 1 receptors in the rat brainstem and nodose ganglia and characterised the binding profile of [α 33P]dATP. Binding of [α 33P]dATP was fully displaceable by adenosine 5′-triphosphate (ATP), and was found on both human and rat nodose ganglia, and throughout the rat brainstem, including the nucleus tractus solitarius and ventrolateral medulla. [α 33P]dATP binding in the human nodose ganglia was significantly displaced by both 2-methylthio ATP and α,β-methylene ATP, but not by uridine 5′-triphosphate, pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid, 8,8′-(carbonylbis(imino-4,1-phenylenecarbonylimino-4,1-phenylenecarbonylimino))bis(1,3,5-naphtalenetrisulfonic) acid (NF279) or N-ethylcarboxamidoadenosine. [α 33P]dATP binding in the rat nodose ganglia and brainstem was significantly displaced by only 2-methylthio ATP, suggesting that [α 33P]dATP is binding to P2Y receptors in the rat. Binding of [α 33P]dATP was also significantly displaced by α,β-methylene adenosine 5′-diphosphate, suggesting a component of the binding is to endogenous ecto-5′-nucleotidase, however, almost all binding could be displaced by a combination of receptor agonists (2-methylthio ATP, uridine 5′-triphosphate and α,β-methylene ATP), suggesting preferential binding to receptors. Immunoreactivity to P2Y 1 receptor (P2Y 1-IR) exhibited similar distribution patterns to [α 33P]dATP binding, with a clear topographic profile. Particularly dense P2Y 1-IR labeling was evident in cells and fibres of the dorsal vagal complex. Immunolabeling was also present in the dorsal motor nucleus of the vagus and nucleus ambiguus, indicating the possibility of P2Y 1 receptors on vagal efferents. Unilateral vagal ligation was also performed to examine the transport of P2Y 1 receptor, using both immunohistochemistry and [α 33P]dATP autoradiography. Accumulations of both P2Y 1-IR and [α 33P]dATP binding were apparent adjacent to both ligatures, suggesting bi-directional transport of P2Y 1 receptors along the rat vagus nerve. This current study represents the first description of P2Y 1 receptor distribution within the rodent brainstem and nodose ganglion and also characterises [α 33P]dATP binding to P2Y receptors.