Calcium-binding proteins in primate cerebellum

Abstract

Single and double antigen localization procedures were used to study the distribution of the calcium-binding proteins calretinin, calbindin and parvalbumin in the cerebellum of the squirrel monkey (Saimiri sciureus). The immunostaining experiments have revealed that each of the three calcium-binding proteins occurred, either alone or in various combinations, in many neuronal types of the monkey cerebellum, including the Purkinje cells. Immunoreactivity for calbindin was detected in virtually all Purkinje cells, whereas immunoreactivity for calretinin and parvalbumin was encountered only in some subpopulations of Purkinje cells. In the vermal region, parvalbumin immunostaining appeared in the form of typical weak and strong alternating parasagittal bands. Calretinin immunoreactivity was found in virtually all neurons and fiber systems related to the granular layer, including the monodendritic cells, the granule cells and their parallel fibers, the Golgi and Lugaro cells and the mossy fibers. The Golgi cells also displayed calbindin and parvalbumin immunoreactivity. Parvalbumin was found to labeled both the climbing and mossy fibers, as well as the basket and stellate cells lying in the molecular layer. These results reveal that virtually all the different neuronal types in the primate cerebellum contain at least one of three calcium-binding proteins investigated in the present study. Furthermore, calretinin appears to be a particularly reliable molecular maker for all the neuronal elements associated with the granular layer in the primate cerebellum.

Introduction

The cerebellum is concerned with the coordination of somatic motor activity, the regulation of motor tone, and mechanisms that influence and maintain equilibrium. Structurally, the cerebellum consists of: (a) a superficial gray mantle, the cerebellar cortex; (b) an internal white mass, the medullary substance; and (c) four pairs of intrinsic nuclei embedded in the white matter. The cerebellar cortex is uniformly structured in all parts and expands across the midline without evidence of a median raphe. The cortex is composed of three well defined layers containing different types of neurons. These layers from the surface are: (a) the molecular layer; (b) the Purkinje cell layer; and (c) the granular layer (Ito, 1984, Parent, 1996).

In order to exert its multifarious roles, the cerebellum uses complex neural networks composed of specific subsets of chemospecific elements. In the present study, attempts were made to identify immunohistochemically cerebellar neurons that contain calcium-binding proteins, which are known to play a crucial role in the maintenance of intracellular calcium homeostasis. Our attention was focused on three calcium-binding proteins characterized structurally by EF-hand motifs, namely calretinin (CR), calbindin-D28k (CB) and parvalbumin (PV). These three proteins have been extensively used as neuronal markers of specific neuronal populations in the central nervous system (Celio, 1990, Baimbridge et al., 1992, Résibois and Rogers, 1992, Rogers and Résibois, 1992, Seto-Ohshima, 1994, Parent et al., 1996). Information about the localization of calcium-binding proteins in the cerebellum is available for rats and chicks (Rogers, 1989a, Rogers, 1989b), but nothing is known of the neuronal distribution of these proteins in the cerebellum of monkeys. We therefore thought of interest to undertake a detailed study of the cellular localization and degree of colocalization of CR, CB or PV in the cerebellum of the squirrel monkey in the hope to shed a new light on the complex neurochemical organization of the cerebellum, as well as on the role of calcium-binding proteins in the brain.