Na/Ca exchanger isoforms in developing rat lens

Abstract

Objective: Although Ca homeostasis is essential to the maintenance of lens transparency and the control of lens fiber cell differentiation, the molecular mechanism of Ca regulation remains unclear. Na/Ca exchanger (NCX) catalyzed reversible exchange of Na for Ca across the plasma membrane. To reveal the role of NCX in the control of lens Ca homeostasis, we characterized NCX isoforms expressed in developing rat lens. Methods: The lens from neonatal (post natal day 5, 7 and 9) and adult Wistar rats were homogenized in the lysis buffer containing 20mM HEPES (pH 7.4), 150mM NaCl, 1% sodium deoxycholate, 1% Triton X-100, 0.1% SDS, 2μg/ml leupeptin, 1μg/ml aprotinin, 200μM phenylmethylsulfonyl fluoride, and 200mM benzamidine hydrochroride. The lysates were centrifuged and resultant supernatants were subjected to SDS-PAGE on 8.5% gel and then to immunoblotting with anti-NCX1, 2, and 3 antibody. Results and Discussion: Anti-NCX1 antibody recognized the molecule of 120KDa, which is mature NCX1, in both neonatal and adult rat lens, while Anti-NCX2 and 3 antibodies did not detect any band. In rat lens, including developmental period, NCX1 acts a role as a Ca extrusion system to modulate the intracellular Ca concentration. To establish the causal relationship between NCX activity and lens transparency, we are trying a characterization and biochemical analysis of NCX1 using the lens of experimental rat cataract models. [J Physiol Sci. 2007;57 Suppl:S221]