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. 2015 Sep;146(3):205–219. doi: 10.1085/jgp.201511383

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© 2015 López Soto et al.

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Figure 3. — Ca_V2.2 inhibition by constitutive and ghrelin-dependent GHSR1a activity is signaled by G_i/o and G_q proteins, respectively. (A) Time course, representative traces, and averaged I_Ca inhibition by 500 nM ghrelin in HEK 293T cells transfected with Ca_V2.2, Ca_Vα₂δ₁, Ca_Vβ₃, and GHSR1a-A204E in control conditions or preincubated with 500 ng/ml ChTx or 500 ng/ml PTx, or cotransfected with a G_qDN. (B) Representative traces and averaged I_Ca in HEK 293T cells expressing Ca_V2.2, Ca_Vα₂δ₁, Ca_Vβ₃, and GHSR1a or GHSR1a-A204E in control or preincubated with 500 ng/ml ChTx or 500 ng/ml PTx, or cotransfected with G_qDN. (C) Representative traces and averaged I_Ca inhibition by 500 nM ghrelin in HEK 293T cells transfected with Ca_V2.2, Ca_Vα₂δ₁, Ca_Vβ₃, and GHSR1a preincubated with 500 ng/ml PTx. (D) Representative I_Ca in HEK 293T cells cotransfected with Ca_V2.2, Ca_Vα₂δ₁, GHSR1a-A204E, and Ca_Vβ₃ or Ca_Vβ_2a without (control) or with (+ghrelin) 500 nM ghrelin without or with (+pp) an 80-mV prepulse application (left), and averaged percentage of I_Ca inhibition by 500 nM ghrelin in cells expressing Ca_V2.2, Ca_Vα₂δ₁, and GHSR1a-A204E with the coexpression of Ca_Vβ₃ or Ca_Vβ_2a and MAS-GRK2-ct and prepulse application (+pp; right). (E) Representative I_Ca traces from cells cotransfected with Ca_V2.2, Ca_Vα₂δ₁, Ca_Vβ₃, and GHSR1a or GHSR1a-A204E with (+pp) or without (Control) the application of an 80-mV prepulse (left) and averaged I_Ca from HEK 293T cells expressing Ca_V2.2, Ca_Vα₂δ₁, GHSR1a, or GHSR1a-A204E, with the coexpression of Ca_Vβ₃ or Ca_Vβ_2a and MAS-GRK2ct and an 80-mV prepulse application (+pp; right). ANOVA with Dunnett’s (A and B) or Tukey’s post-test (D), and two-sample Student’s t test (E). *, P < 0.05. Error bars represent mean ± SE.