Whole-cell recording of the robust nucleus of the arcopallium neurons in the adult zebra finch

BACKGROUND： Electrophysiological properties of the song nucleus have been revealed using conventional techniques, such as intracellular and extracellular recording. Research concerning the neuronal activation properties and regulations of the song system at the cellular and ion channel level may help reveal the neural mechanism of song learning. OBJECTIVE： To perform whole-cell recording of robust nucleus of the arcopallium （RA） neurons in brain slices from adult zebra finches （Taeniopygia guttata） and observe the action potential, sodium/potassium current and the spontaneous postsynaptic current of RA neurons. DESIGN, TIME AND SETTING： Self-controlled, neuroelectrophysiological experiment. The study was performed at the Neurophysiology Laboratory of South China Normal University from April to September 2008. MATERIALS： Flaming/Brown puller P-97 was purchased from Sutter Ins, USA; Axopatch 700B amplifier and Digidata 1332A converter were purchased from Axon Instrument, USA; pClamp software was provided by Axon Instrument, USA. METHODS： RA neurons were acutely isolated from 24 healthy male zebra finches. The action potential, voltage-gate sodium/potassium current and spontaneous postsynaptic current were recorded by whole-cell recording technology. Data were analyzed by pClamp software. MAIN OUTCOME MEASURES： The amplitude and frequency of the action potential, and the amplitude of the voltage-dependent and spontaneous postsynaptic currents, were measured. RESULTS： （1） Testing of action potential： Cells exhibited a stable current-voltage relationship following a series of hyperpolarization stepped currents, and an action potential was triggered by the spike threshold. All the recorded cells displayed repetitive firing following depolarizing current injection, with a frequency beyond 100 Hz. （2） Testing of voltage-gate currents： The inward and outward whole-cell currents were observed after a series of depolarizing voltage steps. The inward current disappeared following the application of tetrodotoxin and the outward current was significantly inhibited by application of 4-aminopyfidione and tetraethylammonium chloride. （3） Testing of spontaneous postsynaptic current： The majority of recorded cells exhibited an inward synaptic current when the membrane potential was maintained at -60 mV, with some cells exhibiting a robustly outward current when the membrane potential was maintained at -30 mV. Tetrodotoxin was unable to affect the spontaneous postsynaptic current. Following application of bicuculline [y-aminobutyric acid （A） receptor antagonist] and high concentration kynurenic acid （ionotropic glutamate receptor antagonist）, the inward and outward currents were completely inhibited. CONCLUSION： Under these experimental conditions, the action potential, sodium/potassium current and spontaneous postsynaptic current were recorded successfully in RA neurons. This indicates that the cells preserved relatively intact synaptic connections and normal physiological activity, which is required for investigating ion channels. The inward and outward whole-cell currents were sodium and potassium currents, respectively. The postsynaptic y-aminobutyric acid （A） receptors and ionotropic glutamate receptors contributed to the spontaneous postsynaptic current.