Voltage-clamp analysis of neurons within deep layers of the brain

Single electrode whole cell current- and voltage-clamp techniques in conjunction with intra- and extracellular phoresis and extracellular application of pharmacological agents were applied to study neurons in deep layers of the brainstem of anesthetized, paralyzed and artificially ventilated cats. We compared slow rhythmic changes and stimulus-evoked postsynaptic current and voltage responses of neurons as they were recorded with fine-tipped microelectrodes filled with 2-3 M 'microelectrode solutions' or with 0.3 M 'patch solutions', or with patch electrodes. The experimental data were then compared with the effects of somatic and dendritic conductance changes simulated in a cell model. A new method was introduced for alternating current and voltage-clamp measurements performed at 300 Hz, which provided quasi-simultaneous measurements of slow changes of spontaneous synaptic currents and potentials. During current or voltage clamp, chemicals which affect voltage- and receptor-controlled conductances were ionophoresed intracellularly through single or theta-type glass electrodes. We show examples of activation of low-voltage activated Ca2+ responses after blockade of Na+ currents by intracellular QX 314 and K+ currents by intracellular Cs+ injections in addition to Sp-cAMPs to activate protein kinase A. TEA, NMDA and GABA were used to demonstrate the effectiveness of extracellular application of drugs through multibarrel electrodes or local application through a 'bath'. The various tests demonstrated that single electrode whole cell current- and voltage-clamp methods, in combination with various techniques for drug application, can be well applied to study the biophysical properties and pharmacological sensitivities of neurons embedded in in vivo networks within deep layers of the brain.