ABSTRACT
Whole-cell patch-clamp technique was employed to record chloride ionic current IGABA evoked by fast (600 msec) application of GABA to hippocampal pyramidal neurons and cerebellar Purkinje cells isolated from rat brain. GABA solution in the application pipette was either neutral (pH 7.4) or acidic (pH 7.0 or 6.0). Application of protons to neurons causes a rapid, reversible, and dose-dependent decrease in the amplitude of IGABA; the effect was more pronounced on hippocampal neurons (carrying both synaptic and extrasynaptic GABAA receptors) than in cerebellar Purkinje cells (predominantly equipped with synaptic GABAA receptors). In hippocampal neurons, pharmacological isolation of extrasynaptic component from total IGABA was performed with GABAA receptor antagonist gabazine (50 nM). The extrasynaptic component of IGABA was stronger blocked by protons than total IGABA. It was concluded that acidic medium produced more potent blocking effect on extrasynaptic GABAA receptors than on synaptic ones.
Subject(s)
Evoked Potentials/drug effects , Protons , Purkinje Cells/drug effects , Pyramidal Cells/drug effects , Receptors, GABA-A/physiology , gamma-Aminobutyric Acid/pharmacology , Animals , CA3 Region, Hippocampal/cytology , CA3 Region, Hippocampal/drug effects , CA3 Region, Hippocampal/physiology , Cerebellum/cytology , Cerebellum/drug effects , Cerebellum/physiology , Dose-Response Relationship, Drug , Evoked Potentials/physiology , GABA Antagonists/pharmacology , Hydrogen-Ion Concentration , Patch-Clamp Techniques , Primary Cell Culture , Protein Subunits/physiology , Purkinje Cells/cytology , Purkinje Cells/physiology , Pyramidal Cells/cytology , Pyramidal Cells/physiology , Pyridazines/pharmacology , Rats , Rats, WistarABSTRACT
Taurine is an important endogenous agonist of glycine receptors (GlyR). Using the patchclamp technique, we measured chloride current induced by a short (600 msec) application of taurine (ITau) on isolated rat pyramidal neurons. pH of taurine solution in the applicator pipette was neutral (7.4) or acidic (7.0-5.0). Application of protons to a neuron causes a dosedependent decrease in the peak amplitude and acceleration of ITau desensitization. Addition of 100 nM ß-amyloid peptide (Aß) to the perfusate caused acceleration of ITau desensitization. The effects of Aß and H+ on the rate of ITau desensitization were not additive. In addition, Aß attenuated the effect of H+ on the peak amplitude of ITau. We also studied the effect of protons on the chloride current caused by activation of GABA receptors. In contrast to H+ effects on GlyR, Aß did not modulate the effects of H+ on GABA receptors.
Subject(s)
Amyloid beta-Peptides/pharmacology , Chlorides/metabolism , Hippocampus/cytology , Protons , Pyramidal Cells/drug effects , Pyramidal Cells/metabolism , Taurine/pharmacology , Animals , Hydrogen-Ion Concentration , Patch-Clamp Techniques , RatsABSTRACT
Voltage clamp and concentration-jump methods were employed to examine the effects of endogenous neuropeptide cycloprolylglycine on GABA-activated ionic currents in isolated cerebellar Purkinje cells. In the concentration range of 0.1-10.0 µM, short-term (600 msec) external application of cycloprolylglycine against the background of GABA-evoked current produced no effect on its amplitude. In contrast, application of 1 µM cycloprolylglycine increased current up to 177±15% control level. The development of potentiating effect and return to the control level of ionic current were slow, which was indicative of possible implication of second messenger systems in these processes. Functional augmentation of GABAA receptors under the action of cycloprolylglycine can underlie the established neuroprotective and anxiolytic effects of this endogenous dipeptide.
Subject(s)
Peptides, Cyclic/pharmacology , Purkinje Cells/drug effects , Purkinje Cells/metabolism , Receptors, GABA-A/metabolism , Animals , Cells, Cultured , GABA Agonists/pharmacology , Humans , Patch-Clamp Techniques , Peptides, Cyclic/metabolism , RatsABSTRACT
The concentration-clamp experiments with neurons isolated from the rat brain showed that nootropic and neuroprotective drug Semax added to perfusion solution at concentration of 1 µM augmented the amplitude of GABA-activated ionic currents in cerebellum Purkinje cells by 147±13%. In addition, Semax in perfusion solution (0.1 and 1 µM) diminished the amplitude of glycine-activated chloride currents in hippocampal pyramidal neurons down to 68 and 43% control level, respectively. Both potentiating and inhibitory effects developed slowly, and they were poorly reversible, which indicated a probable implication of second messengers in the observed phenomena. Semax accelerated the falling edge of glycine-activated current both after a short-term co-application with agonist and after addition of this peptide into perfusion solution.
Subject(s)
Adrenocorticotropic Hormone/analogs & derivatives , Cerebellum/cytology , Glycine/metabolism , Hippocampus/cytology , Neurons/drug effects , Neurons/metabolism , Peptide Fragments/pharmacology , gamma-Aminobutyric Acid/metabolism , Adrenocorticotropic Hormone/pharmacology , Animals , Cells, Cultured , RatsABSTRACT
The effects of donepezil on voltage-dependent Ca(2+)- and low-threshold K(+)-current were studied on isolated molluscan neurons using two-electrode voltage clamp technique. Donepezil reduced the amplitude of voltage-dependent Ca(2+)-current (IC(50)=7.9 microM) and shifted the current-voltage relationships toward hyperpolarization. Donepezil in low concentration (5 microM) increased, while in higher concentrations (>or=10 microM) decreased the low-threshold K(+)-current.
Subject(s)
Cholinesterase Inhibitors/pharmacology , Indans/pharmacology , Ion Channels/physiology , Neurons/physiology , Piperidines/pharmacology , Animals , Calcium Channels/drug effects , Calcium Channels/physiology , Cell Membrane/drug effects , Cell Membrane/physiology , Donepezil , Helix, Snails , Ion Channels/drug effects , Membrane Potentials/drug effects , Membrane Potentials/physiology , Microelectrodes , Neurons/drug effects , Patch-Clamp TechniquesSubject(s)
8-Bromo Cyclic Adenosine Monophosphate/metabolism , Chloride Channels/metabolism , Glutamic Acid/chemistry , Neurons/drug effects , Animals , Chlorides/chemistry , Cyclic AMP/metabolism , Electrodes , Electrophysiology , Hippocampus/metabolism , Membrane Potentials , Mollusca , Neurons/metabolism , Patch-Clamp Techniques , Software , Synaptic Transmission , Time FactorsABSTRACT
Two-microelectrode voltage clamping experiments were performed on isolated snail neurons to measure the Ca2+-dependent. potential-dependent K+ current (I(C)), with assessment of the effects of penetrating cGMP analogs on this current - dibutyryl cGMP (dcGMP) and 8-Br-cGMP. Both of these penetrating cGMP analogs rapidly and reversibly decreased the amplitude of I(C). cGMP analogs produced no shifts in the volt-ampere characteristics of the efflux current along the voltage axis. dcGMP and 8-Br-cGMP had no effect on the influx Ca2+ current. The non-specific protein kinase inhibitor H-8 decreased or had no effect on I(C) in different cells. The effects of both dcGMP and 8-Br-cGMP persisted in the presence of H-8. Decreases in I(C) in the presence of cGMP analogs also persisted in the presence of the protein phosphatase inhibitor okadaic acid. These results lead to the conclusion that decreased conductivity of Ca2+-dependent K+ channels occurring in response to cGMP is not associated with phosphorylation.
