GABA(A) receptors on calbindin-immunoreactive myenteric neurons of guinea pig intestine

These studies were carried out to characterize the properties of gamma-amin obutyric acid(A) (GABA(A)) receptors on guinea pig intestinal myenteric neu rons maintained in primary culture. In addition, the type of neuron express ing GABA(A) receptors was identified using immunohistochemical methods. Who le-cell patch clamp recordings of currents elicited by GABA and acetylcholi ne (ACh) were obtained using pipettes containing Neurobiotin(R). After elec trophysiological studies, neurons were processed for localization of calbin din-D28K-immunoreactivity (calbindin-ir). GABA (1 mM) and ACh (3 mM) caused inward currents in most cells tested. GABA currents were mimicked by musci mol (1-300 mu M) and were blocked by bicuculline (10 mu M) indicating that GABA was acting at GABA(A) receptors. GABA currents were associated with a conductance increase and a linear current/voltage relationship with a rever sal potential of 1 +/- 1 mV (n = 5). Pentobarbital (PBI 3-1000 mu M) and di azepam (DZP, 0.01-10 mu M) potentiated GABA-induced currents. A maximum con centration of DZP (1 mu M) increased GABA-induced currents 3.1 +/- 0.3 time s while PB (1000 mu M) increased GABA currents by 11 +/- 2 times. In outsid e-out patches, the amplitude of GABA-activated single-channel currents was linearly related to membrane potential with a single-channel conductance of 28.5 +/- 0.5 pS (n = 10). PB and DZP increased thr open probability of GAB A-induced single-channel currents. Neurons containing calbindin-ir were lar ge, were isolated from other neurons and had GABA current amplitudes of -3. 4 +/- 0.3 nA (n = 48). Neurons with weak or absent calbindin-ir were smalle r. were localized in clusters of cells and had GABA-induced current amplitu des of -0.6 +/- 0.1 nA (n = 20). ACh-induced currents were smaller, in calb inclin-ir neurons (-0.7 +/- 0.1 nA) compared to weakly calbindin-ir neurons (-1.4 +/- 0.1 nA). These results indicate that myenteric calbindin-ir neur ons express a high density of GABA(A) receptors. Cell size and location all ow visual identification of neurons likely to contain calbindin-ir permitti ng targeted studies of the properties of these neurons. (C) 2000 Elsevier s cience B.V. All rights reserved.