Selective inhibition of high voltage-activated L-type and Q-type Ca2+ currents by serotonin in rat melanotrophs

1. Whole-cell Ca2+ currents (I-Ca) from cultured rat melanotrophs were identified by their sensitivity to Ca2+ channel blockers, and their modulation by serotonin (5-HT) was studied. All cells displayed high voltage-activated (HVA; > -30 mV) Ca2+ currents. A low voltage-activated (LVA; > -60 mV) Ca2+ current was detected in 92% of the cells. 2. The whole-cell I-Ca was insensitive to omega-conotoxin GVIA (0.5-1 mu M) indicating the absence of N-type Ca2+ channels. 3. At a holding potential (V-h) of -70 mV, the L-type channel blocker nifedipine reduced I-Ca in a dose-dependent manner with a half-maximal effective concentration (IC50) of 28 nM. The L-type current represented 39% of the total I-Ca. 4. omega-Agatoxin IVA (omega-Aga IVA) produced a biphasic dose-dependent inhibition of I-Ca, with IC50 values of 0.4 and 91 nM, indicating the presence of P-type and Q-type Ca2+ channels. which accounted respectively for 16 and 45% of the total I-Ca. The P-type current was also blocked by synthetic funnel-web spider toxin (sFTX 3.3; 1-10 mu M) and was present only in a subpopulation (60-70%) of cells. 5. All cells possessed a Ca2+ current which was resistant to nifedipine (10 mu M) and omega-Aga IVA (50 nM). This current was not affected by Ni2+ (40 mu M) but was abolished by a low concentration of Cd2+ (10 mu M) and by omega-conotoxin MVIIC (1 mu M) indicating that it was a Q-type Ca2+ current. 6. 5-HT (10 mu M) inhibited the whole-cell I-Ca in 70% of the cells tested (n = 120) by activating 5-HT1A and 5-HT2C receptors. 5-HT produced either a kinetic slowing of the activation phase (37% of the cells) or a scaling down (14% of the cells) of I-Ca. In the majority of cells (49%) both types of inhibition were found to coexist. 7. The effects of 5-HT were voltage dependent, rendered irreversible when GTP-gamma-S (30 mu M) was present in the pipette solution and abolished by pretreatment of the cells with pertussis toxin (PTX; 150 ng ml(-1), 18 h). 8. Low concentrations of omega-Aga IVA (20 nM), which blocked mainly P-type channels, did not reduce the effect of 5-HT on I-Ca. The scaling down effect of 5-HT on I-Ca was eliminated in the presence of nifedipine (10 mu M) and the kinetic slowing effect of 5-HT persisted after blockade of L- and P type channels but was abolished by omega-conotoxin MVIIC (1 mu M). 9. We conclude that rat melanotrophs possess functional L-, P- and Q-type Ca2+ channels and that 5-HT inhibits selectively L-type and Q-type Ca2+ currents with different modalities. These effects are voltage dependent and mediated by a PTX-sensitive G-protein.