Functional P2X7 Receptor Expression in the Magnocellular Neurons of the Hypothalamic Neurohypophysial System (HNS)

Abstract

Our laboratory has presented evidence for P2x7 receptors (P2x7R) in isolated terminals of the neurohypophysis (NH) (Cuadra et al., 2010, Biophys. J.97: 1474). In this study we further examined P2x7R immunoreactivity (IR) and function. A stereo-specific ecto-P2x7R antibody was used for IR studies (Kim el al., 2001, JBC, 276:23262) in the rat HNS: supraoptic nucleus (SON), paraventricular nucleus (PVN) and NH. We found limited P2x7R IR in the SON and the PVN, which was absent from the somata of OT- and AVP-neurons. P2x7R IR was co-localized in some cell bodies containing the glial specific marker, GFAP. In contrast, P2x7R IR was abundantly seen on the membrane of NH terminals (NHT), with dense IR in terminals lining the capillary borders. This was confirmed in isolated NHT that showed IR puncta on the membranes of both AVP- and OT-containing terminals. Initial results using ratiometric calcium imaging with Fura-2 in isolated NHT showed differential [Ca2+]iresponses to 100 μM vs. 1 mM ATP; some NHT responded to both doses while others responded only to 1 mM ATP. A similar distribution in response was observed with ATP generated ion currents during patch clamp recordings. NHT responded with either high sensitivity (HS) or low sensitivity (LS) to ATP. The HS group corresponded to AVP-NHT, which is reported to express P2x2R, P2x3R, P2x4R and P2x7R (Knott et al., 2005, Pfluger Arch, 405:381). In contrast, the LS group corresponded to OT-NHT, which is consistent with a P2x7R response. Together these data suggest that in the HNS P2x7R is expressed chiefly in the AVP- and OT-secreting terminals of magnocellular neurons. (Supported by NIH grant NS29470 to JRL)