Cloues, Robin Kimberly; (1994) Atp-activated channels in rat superior cervical ganglion neurones and their modulation by extracellular zinc. Doctoral thesis (Ph.D.), University College London (United Kingdom).

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Abstract

It has recently been shown that adenosine 5'-triphosphate (ATP) can act as a fast excitatory transmitter at neuro-neuronal synapses in both the central and the peripheral nervous systems. I have examined the ATP-activated inward current (IATP) cultured rat superior cervical ganglion (SCG) neurones and its modulation by extracellular zinc ions (Zn2+). ATP activated a non-specific cation conductance and caused a transient rise in intracellular Ca2+ which was dependent on extracellular Ca2+. The current was activated specifically by ATP and was reversibly blocked by the P2-purinoceptor antagonist, suramin. Low concentrations of extracellular Zn2+ rapidly and reversibly potentiated both IATP and the intracellular Ca2+ rise. Higher concentrations of Zn2+ reduced and prolonged the current. Other divalent cations mimicked the effect of Zn2+ with an order of potency of: Cu2+ [greater-than] Zn2+ [greater-than] Ni2+ [greater-than] Cd2+ [greater-than] Co2+ [greater-than] Mn2+. The potentiation by Zn2+ was dependent on the concentration of agonist; Zn2+ increased the apparent affinity of the receptor for ATP without potentiating the maximum response. Single channels activated by ATP, and reversibly blocked by suramin, were recorded using excised outside-out patches. The channels were small (conductance at -80 mV = 11 pS). Both single channel conductance and opening probability increased with hyperpolarization. Low concentrations of Zn2+ significantly increased the frequency of ATP-evoked channel opening and burst duration without altering the unitary conductance. Higher concentrations of Zn2+ further increased channel burst duration but also decreased unitary current amplitude. These results are consistent with two sites of action for Zn2+: a positively acting allosteric site which enhances macroscopic current amplitude and a site, possibly within the pore, which blocks conductance through the channel. In conclusion, the P2x-purinoceptor in rat SCG neurones is allosterically regulated by changes in concentration of extracellular Zn2+, increasing both IATP amplitude and ATP-evoked changes in intracellular Ca2+.

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