Presynaptic mechanism of action of 4-aminopyridine: changes in intracellular free Ca2+ concentration and its relationship to B-50 (GAP-43) phosphorylation.
Recently we have shown that 4-aminopyridine (4-AP), a drug known to enhance transmitter release, stimulates the phosphorylation of the protein kinase C substrate B-50 (GAP-43) in rat brain synaptosomes and that this effect is dependent on the presence of extracellular Ca2+. Hence, we were interested in the relationship between changes induced by 4-AP in the intracellular free Ca2+ concentration ([Ca2+]i) and B-50 phosphorylation in synaptosomes. 4-AP (100 microM) elevates the [Ca2+]i (as determined with fura-2) to approximately the same extent as depolarization with 30 mM K+ (from an initial resting level of 240 nM to approximately 480 nM after treatment). However, the underlying mechanisms appear to be different: In the presence of 4-AP, depolarization with K+ still evoked an increase in [Ca2+]i, which was additive to the elevation caused by 4-AP. Several Ca2+ channel antagonists (CdCl2, LaCl3, and diphenylhydantoin) inhibited the increase in B-50 phosphorylation by 4-AP. It is interesting that the increase in [Ca2+]i and the increase in B-50 phosphorylation by 4-AP were attenuated by tetrodotoxin, a finding pointing to a possible involvement of Na+ channels in this action. These results suggest that 4-AP (indirectly) stimulates both Ca2+ influx and B-50 phosphorylation through voltage-dependent channels by a mechanism dependent on Na+ channel activity.
Heemskerk
F M
FM
Division of Molecular Neurobiology, Rudolf Magnus Institute, University of Utrecht, The Netherlands.
Schrama
L H
LH
Ghijsen
W E
WE
De Graan
P N
PN
Lopes da Silva
F H
FH
Gispen
W H
WH
D015761:4-Aminopyridine / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000502:physiology; D004563:Electrochemistry; D019922:GAP-43 Protein; D007425:Intracellular Membranes / Q000378:metabolism*; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009994:Osmolar Concentration; D010716:Phosphatidylinositols / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D015222:Sodium Channels / Q000502:physiology; D013569:Synapses / Q000502:physiology*; D013574:Synaptosomes / Q000378:metabolism
D015220:Calcium Channels; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D010716:Phosphatidylinositols; D010750:Phosphoproteins; D015222:Sodium Channels; D015761:4-Aminopyridine; D002118:Calcium