MAPPING THE LOCATION OF FUNCTIONAL NICOTINIC AND GAMMA-AMINOBUTYRIC ACID(A) RECEPTORS ON HIPPOCAMPAL-NEURONS

To assess the density and distribution of functional nicotinic acetylc holine receptors (nAChRs) and gamma-aminobutyric acid (GABA) receptors on hippocampal neurons, we have combined infrared videomicroscopy wit h a nanorobotic micromanipulator system and studied the receptor-media ted currents by using the whole-cell patch-clamp technique. Acetylchol ine or GABA was applied by pressure ejection onto a small segment of e ither cell soma or dendrite, and the resulting current was measured in cultured hippocampal neurons by using a whole-cell pipette positioned at the cell soma. Type IA nicotinic currents, sensitive to blockade b y methyllycaconitine (1 nM) and alpha-bungarotoxin and associated with alpha 7-subunit-containing nAChRs, type II currents, sensitive to blo ckade by dihydro-beta-erythroidine (100 nM) subserved by alpha 4 beta 2 nAChRs, and GABA-mediated currents were evoked when the agonists wer e applied to either cell soma or the dendrites. Analysis of the curren t amplitude with respect to the membrane area covered by the applied a gonist provided an estimation of the receptor density along the somato -dendritic axis of the hippocampal neurons. Such analysis revealed: 1) a nonuniform distribution for the receptor types studied; 2) a higher density of nAChR and GABA receptors at the dendrites than at the soma ; and 3) an increasing density for both nAChR subtypes with distance f rom the center of the cell soma, but increasing and then decreasing de nsity for the GABA receptor. Exposure of cultured hippocampal neurons to colchicine (100 nM for 3 days) produced a dramatic reduction in the dendritic branching, and this morphological feature was associated wi th a significant decrease in the receptor density, such an effect bein g more prominent for nAChRs than for GABA receptors. Given the high Ca ++ permeability of nAChRs, the dendritic localization of nAChRs sugges t that they are involved in modulating the synaptic efficacy at the le vel of the dendrites.