Mechanisms intrinsic to 5-HT2B receptor-induced potentiation of NMDA receptor responses in frog motoneurones

1 In the presence of NMDA receptor open-channel blockers [Mg2+; (+)-5-methyl-10,11-dihydro-5H- dibenzo[a, d] cyclohepten-5,10-imine maleate (MK-801); 1-amino-3,5-dimethyladamantane (memantine)] and TTX, high concentrations (30 - 100 muM) of either 5-hydroxytryptamine (5-HT) or alpha-methyl-5- hydroxytryptamine (alpha-Me-5-HT) significantly potentiated NMDA-induced depolarizations of frog spinal cord motoneurones. 2 Potentiation was blocked by LY-53,857 (10 - 30 muM), SB 206553 (10 muM), and SB 204741 (30 muM), but not by spiroxatrine (10 muM), WAY 100,635 (1 - 30 muM), ketanserin (10 muM), RS 102221 (10 muM), or RS 39604 (10 - 20 muM). Therefore, alpha-Me-5-HT's facilitatory effects appear to involve 5-HT2B receptors. 3 These effects were G-protein dependent as they were prevented by prior treatment with guanylyl-5'- imidodiphosphate (GMP-PNP, 100 muM) and H-Arg-Pro-Lys-Pro-Gln-Gln-D-Trp-Phe-D-Trp-D-Trp-Met-NH2 (GP antagonist 2A, 3 - 6 muM), but not by pertussis toxin (PTX, 3 - 6 ng ml(-1), 48 h preincubation). 4 This potentiation was not reduced by protein kinase C inhibition with staurosporine (2.0 muM), U73122 (10 muM) or N-(2-aminoethyl)-5-isoquinolinesulfonamide HCl (H9) (77 muM) or by intracellular Ca2+ depletion with thapsigargin (0.1 muM) (which inhibits Ca2+/ATPase). Exposure of the spinal cord to the L-type Ca2+ channel blockers nifedipine (10 muM), KN-62 (5 muM) or gallopamil (100 muM) eliminated alpha-Me-5-HT's effects. 5 The calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphtalenesulfonamide (W7) (100 muM) diminished the potentiation. However, the calcium/calmodulin-dependent protein kinase II (CaM Kinase II) blocker KN-93 (10 muM) did not block the 5-HT enhancement of the NMDA responses. 6 In summary, activation of 5-HT2B receptors by alpha-Me-5-HT facilitates NMDA-depolarizations of frog motoneurones via a G-protein, a rise in [Ca2+](i) from the entry of extracellular Ca2+ through L-type Ca2+ channels, the binding of Ca2+ to calmodulin and a lessening of the Mg2+-produced open-channel block of the NMDA receptor.