Figure 5. Screening by Western blots for K_V1.1–1.6 proteins in rat brain and cerebral arteries.

A, Western blots showed immunoreactive bands corresponding to K_V1.1, 1.3, 1.4 and 1.6 α-subunits in rat brain (Br, left lanes), but not in rat cerebral arteries (C, right lanes). The apparent molecular mass values of the brain proteins were approximately: 60 kDa (1.1), 90 kDa (1.3), 85 kDa (1.4) and 60 kDa (1.6). B, K_V1.2 was detected in rat brain (Br) and cerebral arteries (C) at a molecular mass of ˜80 kDa. The immunoreactive band in the arterial proteins was eliminated by an antigenic competing peptide (+CP). C, brain (Br) or cerebral artery (C) membrane proteins were prepared in control buffer or were enzymatically treated with PNGase F (+P) before analysis by electrophoresis. Treatment with PNGase F reduced the apparent molecular mass of K_V1.2 in both types of tissue, implying N-glycosylation of the mature protein. D, the K_V1.5 α-subunit was detected as a ˜75 kDa band in cerebral arteries (C), but was absent in the brain. The immunoreactive band was eliminated by a competing peptide (+CP). Western results were verified in 3 (K_V1.1, 1.3, 1.4, 1.6), 16 (K_V1.2), 4 (K_V1.2 PNGase F) and 6 (K_V1.5) different protein preparations isolated from 4–10 rats.