FIGURE 4.

Npas4 protein expression in beta cells depends on the CaMK, Akt, and CaN signaling pathways. MIN6 cells (A, B, E, F, I, and J) were kept in either basal medium alone (Basal) or stimulated with 40 mm KCl (KCl) for 2 h in the presence of inhibitors or vehicle control. Mouse islets (C, D, G, H, K, and L) were treated in KRBH containing 2.8 mm (2.8G) or 25 mm (25G) glucose for 2 h in the presence of inhibitors or vehicle control. A–D, under stimulatory conditions with KCl in MIN6 cells (A and B, n = 4) or high glucose in islets (C and D, n = 3), Npas4 protein was not induced when CaMKs were inhibited with 3 μm KN-93. E–H, 10 μm of the Akt inhibitor Akti-1/2 prevented Npas4 induction in response to KCl in MIN6 cells (E and F, n = 3) and high glucose in islets (G and H, n = 3) whereas mTOR inhibition with 25 nm Rapa treatment did not. I–L, a dose of 10 nm FK-506 significantly reduced the induction of Npas4 protein levels following KCl treatment in MIN6 cells (I and J, n = 3) and high glucose in islets (K and L, n = 4). Error bars represent mean ± S.E., and significance was determined using a two-tailed Student's t test. *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001; N.D., not detectable.