P2X Purinergic Receptor Knockout Mice Reveal Endogenous ATP Modulation of Both Vasopressin and Oxytocin Release from the Intact Neurohypophysis

Abstract

Bursts of action potentials are crucial for neuropeptide release from the Hypothalamic Neurohypophysial System (HNS). The biophysical properties of the ion channels involved in release of these neuropeptides cannot explain the efficacy of such bursting patterns on secretion. We have previously shown that ATP, acting via P2X receptors, potentiates only AVP release from HNS terminals, whereas, its metabolite adenosine, via A1 receptors acting on transient Ca2+ currents, inhibits both AVP and OT secretion. Thus, purinergic feedback-mechanisms have been proposed to explain bursting efficacy at HNS terminals. Therefore, in the present study we have used specific P2X receptor knockout (rKO) mice and purportedly selective P2X receptor antagonists to determine the P2X receptor subtype responsible for endogenous ATP induced potentiation of electrically stimulated neuropeptide release. Intact neurohypophyses (NH) from wild type (WT), P2X3 rKO, P2X2&3 rKO and P2X7 rKO mice were electrically stimulated with four 25 second bursts (3V at 39Hz) separated by 21 second interburst intervals with or without the P2X2 and P2X3 receptor antagonists, suramin or PPADS. These frequencies, number of bursts, and voltages were determined to maximize both AVP and OT release by electrical stimulations. Treatment of WT mouse NH with suramin/PPADS significantly reduced electrically stimulated AVP release. A similar inhibition by suramin was observed in electrically stimulated NH from P2X3 and P2X7 rKO mice but not P2X2&3 rKO mice, indicating that endogenous ATP facilitation of electrically stimulated AVP release is mediated primarily by the activation of the P2X2 receptor. Surprisingly, electrically stimulated OT release from WT, P2X3, P2X2&3 and P2X7 rKO mice was potentiated by suramin, indicating non-purinergic effects by this “selective” antagonist. Nevertheless, these results show that sufficient endogenous ATP is released by bursts of action potentials to act at P2X2 receptors in a positive-feedback mechanism to differentially modulate neuropeptide release from CNS terminals.

Extracellular ATP and other nucleotides—ubiquitous triggers of intercellular messenger release

“…In a later study, it was suggested that co-released ATP potentiates vasopressin release through the activation of P2X2Rs, P2X3Rs, P2X4Rs, and P2X7Rs whereas oxytocin release is mediated only through the P2X7R [100]. Using electrical stimulation of isolated and intact mouse posterior pituitaries, the concept was supported that the co-released endogenous ATP acts as a paracrineautocrine messenger, enhancing vasopressin secretion from neurohypophysial nerve terminals [104].…”

“…Examples include noradrenaline [80] or glutamate [49,50]. Additional examples for enhanced messenger release by autocrine action of ATP include the release of vasopressin [104], gonadotrophins [109,110], prolactin [114], IL-1β [291], IL-6 [353], CXCL8 [142], IL-10, or IL-33 [311]. Signal maintenance or amplification by ATP-induced ATP release may thus be of considerably greater physiological relevance than hitherto anticipated.…”

Taste buds as peripheral chemosensory processors

“…4). Interestingly, such autocrine positive feedback has also been described for synaptic ATP release in the carotid body [67] and at terminals of neurosecretory cells in the posterior pituitary [68]. Thus, what was initially interpreted as being strictly a postsynaptic deficit of sensory fibers (absence of postsynaptic receptors in the P2X2/P2X3 knockout mice), is now understood to include a profound deficit in presynaptic release mechanisms in taste bud cells.…”

P2X receptor channels in endocrine glands

“…Electrophysiological evidence for the existence of P2XR currents in vasopressinergic PP terminals but not in terminals labeled for oxytocin have also been shown . Experiments with knockout mice revealed that sufficient endogenous ATP is released by bursts of action potential to act at P2X2Rs, but not at P2X3R or P2X7R …”