title: Gramicidin perforated patch-clamp technique reveals glycine-gated outward chloride current in dissociated nucleus solitarii neurons of the rat.
abstract: 1. The inhibitory response of exogenously applied glycine was investigated in freshly dissociated rat nucleus tractus solitarii neurons under whole cell configuration using new perforated patch-clamp technique termed "gramicidin perforated patch technique," which maintains intact intracellular Cl- concentrations. 2. Using the gramicidin perforated patch technique, at a holding potential (VH) of -45 mV, glycine induced outward currents in a concentration-dependent manner with a EC50 of 4.0 x 10(-5) M and at a Hill coefficient of 1.5. In contrast, using the nystatin perforated patch technique, glycine induced inward currents at the same VH in a concentration-dependent manner with an EC50 of 4.9 x 10(-5) M and at a Hill coefficient of 1.2. 3. The glycine-induced outward currents were blocked by strychnine in a concentration dependent manner with an IC50 of 2.2 x 10(-8) M. The blockade was competitive. 4. The current-voltage relationship for the 10(-5) M glycine response showed a clear outward rectification. 5. Ten-fold change of extracellular Cl- with a large impermeable anion resulted in a 65 mV shift of the reversal potential of glycine-induced currents (EGly), indicating that the membrane behaves like a Cl- electrode in the presence of glycine. 6. The intracellular Cl- activity calculated from the EGly ranged from 7.3 to 18.2 mM, with a mean value of 13.3 mM. 7. The values of EGly in the individual neurons were significantly negative to the resting membrane potentials, suggesting the existence of active transport of Cl-.
authors.affiliation: Department of Bio-Plasticity, Kyushu University Faculty of Medicine, Fukuoka, Japan.
authors.forename: J S
authors.initials: JS
authors.lastname: Rhee
authors.forename: S
authors.initials: S
authors.lastname: Ebihara
authors.forename: N
authors.initials: N
authors.lastname: Akaike
mesh_terms: D000818:Animals; D018118:Chloride Channels / Q000187:drug effects* / Q000502:physiology; D046508:Culture Techniques; D005260:Female; D005998:Glycine / Q000502:physiology*; D006096:Gramicidin / Q000494:pharmacology*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009433:Neural Inhibition / Q000187:drug effects* / Q000502:physiology; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D017552:Solitary Nucleus / Q000187:drug effects* / Q000502:physiology; D009435:Synaptic Transmission / Q000187:drug effects*
chemical_list: D018118:Chloride Channels; D006096:Gramicidin; D005998:Glycine