Morphological and neurochemical differentiation of large granular layer interneurons in the adult rat cerebellum

The granular layer of the cerebellar cortex consists of densely packed neur onal cells, classified into granule cells and large interneurons. In this s tudy, we provide a comparative survey of large granular layer interneurons in the adult rat cerebellum based on both morphological and neurochemical c riteria. To this end, double immunofluorescence histochemistry was performe d by combining antibodies against the cytoplasmic antigen Rat-303. calretin in, the metabotropic glutamate receptor mGluR2 and somatostatin. Based on R at-303/calretinin double immunohistochemistry, three distinct populations o f large granular layer interneurons could be discerned: cells immunopositiv e for Rat-303, calretinin or both. Rat-303 or calretinin single-labeled cel ls represented Golgi cells and unipolar brush cells, respectively. Rat-303/ calretinin double-labeled cells located just underneath the Purkinje cell l ayer represented Lugaro cells. Morphometrical analysis distinguished two po pulations of Rat-303-positive Golgi cells according to their location: verm is versus hemisphere. Immunostaining for the metabotropic glutamate recepto r mGluR2 combined with Rat-303 or calretinin revealed that the majority of Golgi cells (about 90%) appeared to be mGluR2 positive. Lugaro cells were m GluR2 negative. In addition, a limited population of large polymorphous int erneurons in the depth of the granular layer with morphological features re sembling Golgi cells also displayed Rat-303/calretinin immunoreactivity and were mGluR2 negative. Double immunohistochemistry for Rat-303 and somatost atin revealed three populations of labeled cells in the depth of the granul ar layer. Besides double-labeled Golgi cells, Rat-303 or somatostatin singl e-labeled cells were present. Based on mGluR2/sornatostatin and calretinin/ somatostatin double immunostainings, Rat-303 single-labeled cells were foun d to correspond to Rat-303/calretinin-positive. mGluR2-negative Golgi-like cells, while the identity of somatostatin single-labeled cells remained unc lear. The data presented in this article elaborate previous reports on the morpho logical and neurochemical differentiation of large interneurons in the rat cerebellar granular layer. In addition, they indicate that the current clas sification of these cells into Golgi cells, Lugaro cells and unipolar brush cells does not describe the observed neurochemical heterogeneity. (C) 2001 IBRO. Published by Elsevier Science Ltd. All rights reserved.