GABA(A) receptors on calbindin-immunoreactive myenteric neurons of guinea pig intestine.
These studies were carried out to characterize the properties of gamma-aminobutyric acidA (GABA(A)) receptors on guinea pig intestinal myenteric neurons maintained in primary culture. In addition, the type of neuron expressing GABA(A) receptors was identified using immunohistochemical methods. Whole-cell patch clamp recordings of currents elicited by GABA and acetylcholine (ACh) were obtained using pipettes containing Neurobiotin. After electrophysiological studies, neurons were processed for localization of calbindin-D28K-immunoreactivity (calbindin-ir). GABA (1 mM) and ACh (3 mM) caused inward currents in most cells tested. GABA currents were mimicked by muscimol (1-300 microM) and were blocked by bicuculline (10 microM) 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 reversal potential of 1 +/- 1 mV (n = 5). Pentobarbital (PB, 3-1000 microM) and diazepam (DZP, 0.01-10 microM) potentiated GABA-induced currents. A maximum concentration of DZP (1 microM) increased GABA-induced currents 3.1 +/- 0.3 times while PB (1000 microM) increased GABA currents by 11 +/- 2 times. In outside-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 the open probability of GABA-induced single-channel currents. Neurons containing calbindin-ir were large, 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 smaller, were localized in clusters of cells and had GABA-induced current amplitudes of -0.6 +/- 0.1 nA (n = 20). ACh-induced currents were smaller in calbindin-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 neurons express a high density of GABA(A) receptors. Cell size and location allow visual identification of neurons likely to contain calbindin-ir permitting targeted studies of the properties of these neurons.
Zhou
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Department of Pharmacology and Toxicology, Michigan State University, East Lansing 48824, USA.
Galligan
J J
JJ
D000818:Animals; D064026:Calbindins; D002478:Cells, Cultured; D003975:Diazepam / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D018755:GABA Agonists / Q000494:pharmacology; D018757:GABA Modulators / Q000494:pharmacology; D058785:GABA-A Receptor Agonists; D006168:Guinea Pigs; D007150:Immunohistochemistry; D007421:Intestine, Small / Q000187:drug effects* / Q000294:innervation* / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009118:Muscimol / Q000494:pharmacology; D009197:Myenteric Plexus / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism*; D010424:Pentobarbital / Q000494:pharmacology; D011963:Receptors, GABA-A / Q000187:drug effects* / Q000378:metabolism; D064030:S100 Calcium Binding Protein G / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000378:metabolism / Q000494:pharmacology
D064026:Calbindins; D018755:GABA Agonists; D018757:GABA Modulators; D058785:GABA-A Receptor Agonists; D011963:Receptors, GABA-A; D064030:S100 Calcium Binding Protein G; D009118:Muscimol; D005680:gamma-Aminobutyric Acid; D010424:Pentobarbital; D003975:Diazepam