Development of “Pinceaux” formations and dendritic translocation of climbing fibers during the acquisition of the balance between glutamatergic and γ‐aminobutyric acidergic inputs in developing Purkinje cells

Abstract

The acquisition of the dynamic balance between excitation and inhibition in developing Purkinje cells, necessary for their proper function, is analyzed. Newborn (P0) mouse cerebellum contains glutamatergic (VGLUT2-IR) and gamma-aminobutyric acid (GABA)-ergic (VIAAT-IR) axons. The former prevail and belong to climbing fibers, whereas the latter neither colabel with calbindin-expressing fibers nor belong to axons of the cortical GABAergic interneurons. During the first postnatal week, VIAAT-IR axons in the Purkinje cell neighborhood remains very low, and the first synapses with basket fibers are formed at P7, when climbing fibers have already established dense pericellular nets. The descending basket fibers reach the Purkinje cell axon initial segment by P9, immediately establishing axoaxonic synapses. The pinceaux appear as primitive vortex-like arrangements by P12, and by P20 interbasket fiber septate-like junctions, typical of fully mature pinceaux, are still missing. The climbing fiber's somatodendritic translocation occurs later than expected, after the regression of the multiple innervation, and follows the ascending collaterals of the basket axons, which are apparently the optimal substrate for the proper subcellular targeting of the climbing fibers. These results emphasize that chemical transmission in the axon initial segment precedes the electrical inhibition generated by field effects. In addition, GABAergic Purkinje cells, as opposed to glutamatergic projection neurons in other cortical structures, do not begin to receive their excitation to inhibition balance until the end of the first postnatal week, despite the early presence of potentially functional GABAergic axons that possess the required vesicular transport system.

Lack of Molecular-Anatomical Evidence for GABAergic Influence on Axon Initial Segment of Cerebellar Purkinje Cells by the Pinceau Formation

“…When investigated on serial sections, the thin sheets were found to be continuous with electron-lucent astrocytic processes expressing GLT-1 (open arrowheads), suggesting molecular/structural specialization of astrocytic processes enwrapping the AIS. Thus, consistent with the previous studies (Somogyi and Hamori, 1976;Sotelo, 2008), the surface of the AIS in PCs is mainly covered by astrocytes. This, together with infrequent synaptic contacts, suggests that the AIS of PCs is an unlikely target for GABAergic innervation.…”

“…A similar anatomical configuration was also observed in the AIS of PCs at postnatal day 13 (Fig. 1 F), when the pinceau formation starts to be differentiated (Ango et al, 2004;Sotelo, 2008).…”

“…During early postnatal life, axon collaterals originating from three to seven BCs descend to form GABAergic perisomatic synapses and further embrace the AIS of PCs to construct the pinceau formation, a cone-shaped structure at the base of PC somata (Roman y Cajal, 1911;Gobel, 1971;Sotelo and Llinás, 1972;Palay and Chan-Palay, 1974;Rosina et al, 1999;Gianola et al, 2003;Ango et al, 2004;Sotelo, 2008;Ichikawa et al, 2011). The AIS of PCs is enriched with Na V 1.6, L1-CAM neurofascin, AnkG, and ␤IV spectrin (Zhou et al, 1998;Jenkins and Bennett, 2001) and triggers action potentials (Foust et al, 2010).…”

“…BC axons make direct contact with the AIS, but synaptic contacts are rare compared with the hippocampus and neocortex. Instead, the AIS is mostly covered with astrocytic processes in PCs (Palay, 1974;Somogyi and Hamori, 1976;Bobik et al, 2004;Sotelo, 2008).…”

Differential expression of TrkB isoforms switches climbing fiber‐Purkinje cell synaptogenesis to selective synapse elimination

“…Although, mossy fiber-granule cell-PC circuit activity begins between P3-7 (Arsenio-Nunes and Sotelo, 1985;Crepel, 1974;Scelfo and Strata, 2005), it rapidly increases from P5, when olivary and cerebellar TrkB.FL and phospho-TrkB are decreasing, until P21-30 when granule cells and mossy fibers are mature (Hamori and Somogyi, 1983;Takagishi and Yamamura, 1993). Moreover, GABAergic interneurons first synapse onto PCs from P7 (Sotelo, 2008), when activated phospho-TrkB is falling and only TrkB.T is increasing. Similarly, in the inferior olive, the peak of full length and activated TrkB is at P4-5, in parallel with CF-PC synaptogenesis and neosynaptogenesis, but preceding olivary afferent synaptogenesis between days P7-10 ( Gotow and Sotelo, 1987).…”