Differential maturation of GIRK2-expressing neurons in the mouse cerebellum

The cerebellar cortex is among the brain regions showing the highest expression levels of G-protein-gated inwardly-rectifying potassium (GIRK/Kir3) channels. Despite their critical contribution in modulating neuronal excitability during development and adult, the spatiotemporal expression of specific GIRK subunits in identified cerebellar neuron populations is unresolved. To characterize this onset of expression, we examined the GIRK2 protein expression in mouse cerebellum by western blot, light microscopy immunohistochemistry and immunofluorescence during perinatal development. Using western blots, GIRK2 expression was low at birth but reach its maximum at P5 before decreasing gradually to adult levels. Immunohistochemical localization indicated that GIRK2 is expressed in granule cells from early stages of development. At the embryonic stage, immunofluorescence techniques for the transcription factor Pax6 allowed to demonstrate that GIRK2 is expressed in granule cell precursors. This GIRK2 expression in granule cells continued throughout postnatal development and adulthood. In addition, the expression of Pax2-GFP allowed selective visualization of Golgi cells during pre- and postnatal development. We could not detect co-expression of Pax2-GFP and GIRK2 during prenatal and early postnatal development, but only at post-migratory stages of Golgi cells, once they are morphologically differentiated and located at the granule cell layer. In the adult cerebellum, we performed a detailed characterization on the expression of GIRK2 in different subpopulations of Golgi cells, using metabotropic glutamate receptor 2 (mGlu(2)) and neurogranin as markers, in GlyT2-GFP and GAD67-GFP mice, and showed that GIRK2 is present in at least four morphological and neurochemical non-overlapping populations of Golgi cells. Altogether, these findings shed new light on the developmental regulation of GIRK channels in the cerebellum, and the main expression in granule cells during perinatal development support the idea that GIRK2 may provide a significant route for modulating different aspects of cerebellar development.