Glutamate-immunoreactive climbing fibres in the cerebellar cortex of the rat

Abstract

The climbing fibre system, one of the two main excitatory inputs to the cerebellar cortex, is anatomically and physiologically well characterized, while the nature of its neurotransmitter is still a matter of debate. We wished to determine whether glutamate-immunoreactive profiles with the morphological characteristics of climbing fibres could be found in the rat cerebellar cortex. For this purpose, a monoclonal 'anti-glutamate' antibody has been used in combination with a sensitive postembedding immunoperoxidase method on semi-thin sections or in combination with a postembedding immunogold method on ultrathin sections. At the light microscopic level, climbing fibres appeared as strongly stained fibrous profiles, chains of interconnected varicosities or heavily labelled dots of various sizes, often in close apposition to principal Purkinje cell dendrites. At the electron microscopic level, certain labelled varicosities or more elongated profiles resembling climbing fibre terminals were in synaptic contact with dendritic spines of Purkinje cells. Quantitative analysis of gold particle densities showed that such elements were about three to four times more heavily labelled than their postsynaptic partners. The results obtained in this study demonstrate that at least a subset of climbing fibres and their terminals contain relatively high levels of glutamate-like immunoreactivity and provide additional evidence for a role of glutamate as transmitter in these cerebellar afferents.

Immunolocalization of the mGluR1b Splice Variant of the Metabotropic Glutamate Receptor 1 at Parallel Fiber‐Purkinje Cell Synapses in the Rat Cerebellar Cortex

“…The use of postembedding immunogold methods for electron microscopy has shown that parallel fiber and climbing fiber synaptic terminals contain high levels of the neurotransmitter glutamate (Somogyi et al, 1986;Grandes et al, 1994a). The release of glutamate from these synaptic terminals in the molecular layer results in the activation of glutamate receptors located on the dendritic tree of Purkinje cells.…”

Precise localization of the voltage-gated potassium channel subunits Kv3.1b and Kv3.3 revealed in the molecular layer of the rat cerebellar cortex by a pre-embedding immunogold method

“…Numerous silver-intensified gold particles were associated with membranes of axons, synaptic terminals and dendritic spines. Strong Kv3.1b labeling was detected in presynaptic varicosities of parallel fibers characterized by containing round synaptic vesicles and forming asymmetrical synapses with Purkinje cell dendritic spines (Palay and Chan-Palay 1974;Grandes et al 1994) (Fig. 1a-d).…”

“…2a-f). Kv3.1b and Kv3.3 were not observed in climbing fiber synaptic boutons identified as bulbous varicosities or as more elongated profiles containing numerous round and electron-translucent synaptic vesicles clustered at synaptic junctions (Palay and ChanPalay 1974;Grandes et al 1994). No specific staining was detected in sections processed without primary antiserum or whenever the primary antiserum was substituted by goat serum.…”

Conserved cellular distribution of the glutamate receptors GluA2/3, mGlu1a and mGlu2/3 in isolated cultures of rat cerebellum

“…GluA2/3 is rarely found in non-synaptic membranes (Bergersen et al, 2005) which may lead to speculate that the cellular distribution of GluA2/3 in the cultured cerebellum is at sites of active glutamatergic synapses. Actually, the only remaining glutamatergic presynaptic elements in the molecular layer are the parallel fibres, as the climbing fibres (Grandes et al, 1994b) were cut off during tissue culturing. Also, glutamatergic synapses in the granule cell layer may come from UBCs as they are the sole remaining glutamatergic input in this layer after the loss of the mossy fibre system.…”