Differential Expression and Association of Calcium Channel Alpha(1b) and Beta Subunits During Rat Brain Ontogeny

Calcium functions as an essential second messenger during neuronal development and synapse acquisition. Voltage-dependent calcium channels (VDCC), which are critical to these processes, are heteromultimeric complexes composed of alpha(1), alpha(2)/delta and beta subunits, beta subunits function to direct the VDCC complex to the plasma membrane as well as regulate its channel properties. The importance of beta to neuronal functioning was recently underscored by the identification of a truncated beta 4 isoform in the epileptic mouse lethargic (Ih) (Burgess, D. L., Jones, J. M., Meisler, M. H., and Noebels, J. L. (1997) Cell 88, 385-392). The goal of our study was to investigate the role of individual beta isoforms (beta 1b, beta 2, beta 3, and beta 4) in the assembly of N-type VDCC during rat brain development. By using quantitative Western blot analysis with anti-alpha(1B)-directed antibodies and [I-125-Tyr(22)]omega-conotoxin GVIA (I-125-CTX) radioligand binding assays, we observed that only a small fraction of the total alpha(1B) protein present in embryonic and early postnatal brain expressed high affinity I-125-CTX-binding sites. These results suggested that subsequent maturation of alpha(1B) or its assembly with auxiliary subunits was required to exhibit high affinity I-125-CTX binding. The temporal pattern of expression of beta subunits and their assembly with alpha(1B) indicated a developmental pattern of expression of beta isoforms: beta 1b increased 3-fold from PO to adult, beta 4 increased 10-fold, and both beta 2 and beta 3 expression remained unchanged, As the beta component of N-type VDCC changed during postnatal development, we were able to identify both immature and mature forms of N-type VDCC, At P2, the relative contribution of beta is beta 1b > beta 3 much greater than beta 2, whereas at P14 and adult the distribution is beta 3 > beta 1b = beta 4, Although we observed no beta 4 associated with the alpha(1B) at P2, beta 4 accounted for 14 and 25% of total alpha(1B)/beta subunit complexes in P14 and adult, respectively. Thus, of the beta isoforms analyzed, only the beta 4 was assembled with the rat a,, to form N-type VDCC with a time course that paralleled its level of expression during rat brain development. These results suggest a role for the beta 4 isoform in the assembly and maturation of the N-type VDCC. [References: 78]

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