CALCIUM FLUX THROUGH PREDOMINANTLY INDEPENDENT PURINERGIC ATP AND NICOTINIC ACETYLCHOLINE-RECEPTORS

Ligand-gated nicotinic acetylcholine receptors (nAChRs) and purinergic ATP receptors are often expressed in the same peripheral and central neurons, and ATP and acetylcholine (ACh) are stored together in some s ynaptic vesicles. Evidence has suggested that nAChRs and ATP receptors are not independent and that some agonists strongly cross-activate an d desensitize both receptor types. Rat sympathetic neurons and nAChRs expressed in Xenopus oocytes were studied to determine the significanc e of the interactions caused by the two agonist types. Current amplitu des induced with separate or combined applications of ATP and nicotine are >90% additive and independent. Half of all neurons tested respond ed to either ATP or nicotine but not to both, indicating differences i n the expression of the two receptors. In neurons that expressed both receptor types, the nAChRs were inhibited by the activity-dependent op en-channel blocker chlorisonadmine. If the purinergic and nicotinic re ceptors were significantly dependent and coactivated, then blocking th e ion channels opened by a nicotinic agonist should diminish the curre nt activated by a purinergic agonist. That result was not seen; rather , complete open-channel block of nAChRs with chlorisondamine did not s ignificantly alter the amplitude or kinetics of ATP-induced currents i n the same neurons. Finally, when cloned nAChR subunits were expressed in oocytes, ATP activated only very small currents compared with the current activated by ACh. For the 13 different nAChR subunit combinati ons that were studied, ATP (50-500 mu M) activated a current that rang ed from 0 to 4% of the size of the current activated by 100 mu M ACh. In summary, we find that there is little cross reactivity, and nAChRs and purinergic ATP receptors are predominately independent, acting wit h separable physiological characteristics. Therefore the quantitative Ca2+ flux could be separately determined for nAChRs and ATP receptors. The fraction of total current that is carried by Ca2+ was quantitativ ely determined by simultaneously measuring the whole cell current and the associated change in intracellular Ca2+ with fura-2. For sympathet ic neurons bathed in 2.5 mM Ca2+ at a holding potential of -50 mV, Ca2 + carries 4.8 +/- 0.3% (mean +/- SE) of the inward current through neu ronal nAChRs and 6.5 +/- 0.1% of the current through purinergic ATP re ceptors. In conclusion, activity-dependent Ca2+ influx through predomi nately independent populations of nAChRs and ATP receptors can produce different intracellular signals at purinergic and cholinergic synapse s.