Organized synapse formation on to Purkinje cell (PC) dendrites by parallel fibers (PFs) and climbing fibers (CFs) is crucial for cerebellar function. In PCs lacking glutamate receptor delta2 (GluRdelta2), PF synapses are reduced in number, numerous free spines emerge, and multiple CF innervation persists to adulthood. In the present study, we conducted anterograde and immunohistochemical labelings to investigate how CFs innervate PC dendrites under weakened synaptogenesis by PFs. In the GluRdelta2 knock-out mouse, CFs were distributed in the molecular layer more closely to the pial surface compared with the wild-type mouse. Serial electron microscopy demonstrated that CFs in the knock-out mouse innervated all spines protruding from proximal dendrites of PCs, as did those in the wild-type mouse. In the knock-out mouse, however, CF innervation extended distally to spiny branchlets, where nearly half of the spines were free of innervation in contrast to complete synapse formation by PFs in the wild-type mouse. Furthermore, from the end point of innervation, CFs aberrantly jumped to form ectopic synapses on adjacent spiny branchlets, whose proximal portions were often innervated by different CFs. Without GluRdelta2, CFs are thus able to expand their territory along and beyond dendritic trees of the target PC, resulting in persistent surplus CFs by innervating the distal dendritic segment. We conclude that GluRdelta2 is essential to restrict CF innervation to the proximal dendritic segment, by which territorized innervation by PFs and CFs is properly structured and the formation of excess CF wiring to adjacent PCs is suppressed.

Synapse formation and functioning are vital for neural network development. Astrocytes ensheathing synapses enable appropriate synapse functioning through glutamate clearance from the synaptic cleft; however, it remains unclear how astrocytic glutamate transporter proteins function at peri-synaptic contact. Here, we report that down syndrome cell adhesion molecule (DSCAM) in Purkinje cells controls climbing fiber (CF) synaptogenesis in the developing cerebellum. Dscam-mutant mice mimicked loss-of-function mutations in the astrocytic glutamate transporter GLAST expressed in Bergmann glia. These mice showed impaired glutamate clearance by the delocalization of GLAST, which complexed with the extracellular domain of DSCAM. Riluzole-induced reduction of free glutamate at the synaptic cleft rescued the impairment of CF synapse formation in Purkinje cell-selective Dscam-deficient mice. DSCAM was required for motor learning, but not gross motor coordination. In conclusion, the intercellular association of synaptic and astrocyte proteins in the tripartite synapse is critical for synapse formation and functioning in neural transmission.