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GABA-mediated Ca²⁺ signalling in developing rat cerebellar Purkinje neurones

Jens Eilers *, Tim D. Plant, Nima Marandi and Arthur Konnerth

1. Cellular responses to GABA_A receptor activation were studied in developing cerebellar Purkinje neurones (PNs) in brain slices obtained from 2- to 22-day-old rats. Two-photon fluorescence imaging of fura-2-loaded cells and perforated-patch recordings were used to monitor intracellular Ca²⁺ transients and to estimate the reversal potential of GABA-induced currents, respectively.
2. During the 1st postnatal week, focal application of GABA or the GABA_A receptor agonist muscimol evoked transient increases in [Ca²⁺]_i in immature PNs. These Ca²⁺ transients were reversibly abolished by the GABA_A receptor antagonist bicuculline and by Ni²⁺, a blocker of voltage-activated Ca²⁺ channels.
3. Perforated-patch recordings were used to measure the reversal potential of GABA-evoked currents (E_GABA) at different stages of development. It was found that E_GABA was about -44 mV at postnatal day 3 (P3), it shifted to gradually more negative values during the 1st week and finally equilibrated at -87 mV at around the end of the 2nd postnatal week. This transition was well described by a sigmoidal function. The largest change in E_GABA was -7 mV day^-1, which occurred at around P6.
4. The transition in GABA-mediated signalling occurs during a period in which striking changes in PN morphology and synaptic connectivity are known to take place. Since such changes were shown to be Ca²⁺ dependent, we propose that GABA-evoked Ca²⁺ signalling is one of the critical determinants for the normal development of cerebellar PNs.

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