Fig. 5. Changes in Cx43–ZO-1 and Cx43-Src associations after exposure to HCH. (A) Lysates from 42GPA9 Sertoli cells that had been incubated with or without 50 µM HCH were immunoprecipitated (Ip) with antibodies against ZO-1, Src or Cx43. Immunoprecipitates were then analyzed by western blotting using antibodies against ZO-1, Src and Cx43. Bands of ZO-1, Src and Cx43 were detected at the predicted molecular masses of 120, 55 and 43 kDa, respectively. In the presence of HCH, levels of ZO-1 associated with Cx43 decreased when lysates were immunoprecipited with either ZO-1 or Cx43. After Cx43 Ip, low but detectable levels of Src were associated with Cx43 in HCH-treated cells, but were totally absent in control cells. (B) Lysates from 42GPA9 Sertoli cells subjected for 1 hour to 50 µM HCH with or without 30 µM PP2 were immunoprecipitated with antibodies against Cx43 or ZO-1. Cx43 and ZO-1 immunoprecipitates were then analyzed by western blotting for P-Src (p-Src) using anti-Src [pY⁴¹⁸] antibody. For each experiment in A and B, a representative blot of three different experiments is shown. Relative optical densities of the bands in arbitrary units are presented in the right panels. Results are the average of three different experiments. ^*P<0.05, ^**P<0.01 significantly different from untreated cells.