P2X(7) is a subtype of ATP-gated channels that is highly expressed in astrocytes, microglia, and other immune cells. Activation of P2X(7) purinoceptors by ATP or 3'-O-(4-benzoyl)-benzoyl ATP (BzATP) induces the formation of cytolytic pores and provokes release of interleukin-1beta from immune cells. We investigated the actions of other endogenous nucleotides on recombinant and microglial P2X(7) receptors using electrophysiology, fluorescence imaging, and interleukin-1beta release measurement. We found that initial application of ADP or AMP to Xenopus oocytes expressing P2X(7) receptors was ineffective. However, when ADP and AMP, but not UTP or adenosine, were applied after a brief exposure to ATP or BzATP, they activated P2X(7) receptors in a dose-dependent manner. Moreover, responses to ADP and AMP were also elicited after exposure to low concentrations of ATP and were recorded several minutes after removal of ATP from the extracellular medium. Whole-cell recordings from mouse microglial cells showed that significant responses to ADP and AMP were elicited only after ATP application. YO-PRO-1 dye uptake imaging revealed that, unlike ATP, prolonged application of ADP or AMP did not cause an opening of large cytolytic pores in mouse microglial cells. Finally, ADP and AMP stimulated the release of interleukin-1beta from ATP-primed mouse and human microglial cells. We conclude that selective sensitization of P2X(7) receptors to ADP and AMP requires priming with ATP. This novel property of P2X(7) leads to activation by ATP metabolites and proinflammatory cytokine release from microglia without cytotoxicity.

Fast chemical communications in the nervous system are mediated by several classes of receptor channels believed to be independent functionally and physically. We show here that concurrent activation of P2X2 ATP-gated channels and 5-HT3 serotonin-gated channels leads to functional interaction and nonadditive currents (47-73% of the predicted sum) in mammalian myenteric neurons as well as in Xenopus oocytes or transfected human embryonic kidney (HEK) 293 cell heterologous systems. We also show that these two cation channels coimmunoprecipitate constitutively and are associated in clusters. In heterologous systems, the inhibitory cross talk between P2X2 and 5-HT3 receptors is disrupted when the intracellular C-terminal domain of the P2X2 receptor subunit is deleted and when minigenes coding for P2X2 or 5-HT3A receptor subunit cytoplasmic domains are overexpressed. Injection of fusion proteins containing the C-terminal domain of P2X2 receptors in myenteric neurons also disrupts the functional interaction between native P2X2 and 5-HT3 receptors. Therefore, activity-dependent intracellular coupling of distinct receptor channels underlies ionotropic cross talks that may significantly contribute to the regulation of neuronal excitability and synaptic plasticity.

Behavioral and neuroendocrine responses underlying systemic osmoregulation are synergistically controlled by osmoreceptors and neuropeptides released within the hypothalamus. Although mechanisms underlying osmoreception are understood, the cellular basis for the integration of osmotic and peptidergic signals remains unknown. Here we show that the excitatory effects of angiotensin II, cholecystokinin and neurotensin on supraoptic neurosecretory neurons are due to the stimulation of the stretch-inactivated cation channels responsible for osmoreception. This molecular convergence underlies the facilitatory effects of neuropeptides on responses to osmotic stimulation and provides a basis for the gating effects of plasma osmolality on the responsiveness of osmoregulatory neurons to peptidergic stimulation.