The effect of endogenous glutamate on neurite outgrowth from cerebellar granule cells in culture was examined. Neurite outgrowth was inhibited by enzymatic removal of endogenous glutamate from the culture medium. The broad-spectrum glutamate receptor antagonist kynurenate also inhibited neurite outgrowth from granule cells in serum-containing and serum-free cultures; the inhibition by kynurenate was reversed by exogenous glutamate. Neurite outgrowth was inhibited to the same extent by the NMDA receptor antagonist APV. These results indicate that endogenous glutamate, possibly released by granule cells themselves, stimulated neurite outgrowth through activation of the NMDA class of glutamate receptors. Activation of NMDA receptors on developing neurons may be an important mechanism for the regulation of neuronal growth and differentiation.

We describe the structure and expression of a mammalian beta‐tubulin isotype (M beta 6) that is weakly expressed in testis but is abundant in developing brain, with transcripts declining to lower levels in the adult brain. The expression of M beta 6 was undetectable in any other mouse tissue examined. A serum specific for this isotype was prepared using a cloned fusion protein as immunogen. M beta 6 is one of five known beta‐tubulin isotypes expressed in brain, and using the anti‐M beta 6 serum along with sera, anti‐M beta 2, anti‐M beta 3/4 and anti‐M beta 5, previously characterized, we have examined the pattern of expression of beta‐tubulin isotypes in rat cerebellum. The isotypes each have characteristic cell‐type specific patterns of localization in cerebellum. M beta 2, M beta 3/4 and M beta 5 are present in both neuronal and non‐neuronal cells, but in contrast M beta 6 was only detectable in neurons in tissue sections and in dissociated cerebellar cell culture. The majority of sequence differences among the beta‐tubulin isotypes lie at the carboxy terminus, the region of beta‐tubulin involved in MAP binding. In the case of M beta 2 and M beta 6, the patterns of expression are similar or identical to the patterns of expression of MAP3 and MAP1A respectively. These results suggest that beta‐tubulin isotypes may contribute to the determination of the specific association of MAPs with microtubules of diverse function. However, the strict subcellular segregation of other MAPs in brain may be determined by other factors.

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Glutamate acting on N-methyl-D-aspartate (NMDA) receptors controls a variety of aspects of neuronal plasticity in the adult and developing brain. This review summarizes its effects on developing cerebellar granule cells. The glutamatergic mossy fibre input to cerebellar granule cells exerts a neurotrophic effect on these cells during development. The investigation of potential neurotrophic agents can be carried out using enriched granule cell cultures. Considerable evidence now indicates that glutamate acting on N-methyl-D-aspartate receptors is an important neurotrophic factor that regulates granule cell development. In culture, neurite growth, differentiation and cell survival are all stimulated by N-methyl-D-aspartate receptor activation. The intracellular pathways involved following Ca2+ entry through the N-methyl-D-aspartate receptor channel are beginning to be elucidated. The cerebellar granule cell culture system may provide an ideal model to investigate the molecular mechanisms involved in long term N-methyl-D-aspartate receptor-mediated changes in neuronal function.

Abstract. The distribution of acetylated ct-tubulin in rat cerebellum was examined and compared with that of total ct-tubulin and tyrosinated ¢t-tubulin. From immunoperoxidase-stained vibratome sections of rat cerebellum it was found that acetylated a-tubulin, detectable with monoclonal 6-11B-l, was preferentially enriched in axons compared with dendrites. Parallel fiber axons, in particular, were labeled with 6-11B-1 yet unstained by an antibody recognizing tyrosinated ¢t-tubulin, indicating that parallel fibers contain ¢t-tubulin that is acetylated and detyrosinated. Axonal microtubules are known to be highly stable and the distribution of acetylated ct-tubulin in other classes of stable microtubules suggests that acetylation and possibly detyrosination may play a role in the maintenance of stable populations of microtubules.