Post-episode depression of GABAergic transmission in spinal neurons of thechick embryo

Whole cell recordings were obtained from ventral horn neurons in spontaneou sly active spinal cords isolated from the chick embryo [embryonic days 10 t o 11 (E10-E11)] to examine the post-episode depression of GABAergic transmi ssion. Spontaneous activity occurred as recurrent, rhythmic episodes approx imately 60 s in duration with 10- to 15-min quiescent inter-episode interva ls. Current-clamp recording revealed that episodes were followed by a trans ient hyperpolarization (7 +/- 1.2 mV, mean +/- SE), which dissipated as a s low (0.5-1 mV/min) depolarization until the next episode. Local application of bicuculline 8 min after an episode hyperpolarized spinal neurons by 6 /- 0.8 mV and increased their input resistance by 13%, suggesting the invol vement of GABAergic transmission. Gramicidin perforated-patch recordings sh owed that the GABAa reversal potential was above rest potential (E-GABAa = -29 +/- 3 mV) and allowed estimation of the physiological intracellular [Cl -] = 50 mM. In whole cell configuration (with physiological electrode [Cl-] ), two distinct types of endogenous GABAergic currents (I-GABAa) were found during the inter-episode interval. The first comprised TTX-resistant, asyn chronous miniature postsynaptic currents (mPSCs), an indicator of quantal G ABA release (up to 42% of total mPSCs). The second (tonic I-GABAa) was comp limentary to the slow membrane depolarization and may arise from persistent activation of extrasynaptic GABAa receptors. We estimate that approximatel y 10 postsynaptic channels are activated by a single quantum of GABA releas e during an mPSC and that about 30 extrasynaptic GABAa channels are require d for generation of the tonic I-GABAa in ventral horn neurons. We investiga ted the post-episode depression of I-GABAa by local application of GABA or isoguvacine (100 muM, for 10-30 s) applied before and after an episode at h olding potentials (V-hold) -60 mV. The amplitude of the evoked I-GABA was c ompared after clamping the cell during the episode at one of three differen t V-hold : -60 mV, below E-GABAa resulting in Cl- efflux; -30 mV, close to E-GABAa with minimal Cl- flux; and 0 mV, above E-GABAa resulting in Cl- inf lux during the episode. The amplitude of the evoked I-GABA changed accordin g to the direction of Cl- flux during the episode: at -60 mV a 41% decrease , at -30 mV a 4% reduction, and at 0 mV a 19% increase. These post-episode changes were accompanied by shifts of E-GABAa of -10, -1.2, and +7 mV, resp ectively. We conclude that redistribution of intracellular [Cl-] during spo ntaneous episodes is likely to be an important postsynaptic mechanism invol ved in the post-episode depression of GABAergic transmission in chick embry o spinal neurons.