Inhibition of N-type calcium channels: the only mechanism by which presynaptic alpha 2-autoreceptors control sympathetic transmitter release.
Alpha 2-Adrenoceptors are known to inhibit voltage-dependent Ca2+ channels located at neuronal cell bodies; the present study investigated whether this or alternative mechanisms, possibly downstream of Ca2+ entry, underlie the presynaptic alpha 2-adrenergic modulation of transmitter release from chick sympathetic neurons. Using chick sympathetic neurons, overflow of previously incorporated [3H]noradrenaline was elicited in the presence of extracellular Ca2+ by electrical pulses, 25 mM K+ or 10 microM nicotine, or by adding Ca2+ to otherwise Ca(2+)-free medium when cells had been made permeable by the calcium ionophore A23187 or by alpha-latrotoxin. Pretreatment of neurons with the N-type Ca2+ channel blocker omega-conotoxin GVIA and application of the alpha 2-adrenergic agonist UK 14304 reduced the overflow elicited by electrical pulses, K+ or nicotine, but not the overflow caused by Ca2+ after permeabilization with alpha-latrotoxin or A23187. In contrast, the L-type Ca2+ channel blocker nitrendipine reduced the overflow due to K+ and nicotine, but not the overflow following electrical stimulation or alpha-latrotoxin- and A23187-permeabilization. The inhibition of electrically evoked overflow by UK 14304 persisted in the presence of nitrendipine and the L-type Ca2+ channel agonist BayK 8644, which per se enhanced overflow. In omega-conotoxin GVIA-treated cultures, electrically evoked overflow was also enhanced by BayK 8644 and almost reached the value obtained in untreated neurons. However, UK 14304 lost its effect under these conditions. Whole-cell recordings of voltage-activated Ca2+ currents corroborated these results: UK 14304 inhibited Ca2+ currents by 33%, nitrendipine caused a 7% reduction, and BayK 8644 increased the currents by 30%. Moreover, the dihydropyridines failed to abolish the inhibition by UK 14304, but pretreatment with omega-conotoxin GVIA, which reduced mean amplitude from 0.95 to 0.23 nA, entirely prevented alpha 2-adrenergic effects. Our results indicate that the alpha 2-autoreceptor-mediated modulation of noradrenaline release from chick sympathetic neurons relies exclusively on the inhibition of omega-conotoxin GVIA-sensitive N-type Ca2+ channels. Mechanisms downstream of these channels and voltage-sensitive Ca2+ channels other than N-type appear not to be important.
Boehm
S
S
Department of Neuropharmacology, University of Vienna, Austria.
Huck
S
S
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D002478:Cells, Cultured; D002642:Chick Embryo; D004563:Electrochemistry; D008564:Membrane Potentials / Q000187:drug effects; D008978:Mollusk Venoms / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects*; D009638:Norepinephrine / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology; D017729:Presynaptic Terminals / Q000187:drug effects*; D018341:Receptors, Adrenergic, alpha-2 / Q000187:drug effects*; D013564:Sympathetic Nervous System / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D020866:omega-Conotoxin GVIA
D002121:Calcium Channel Blockers; D008978:Mollusk Venoms; D010455:Peptides; D018341:Receptors, Adrenergic, alpha-2; D020866:omega-Conotoxin GVIA; D009638:Norepinephrine