Dual action of amitriptyline on NMDA receptors: enhancement of Ca-dependent desensitization and trapping channel block

Although the tricyclic antidepressant amitriptyline (ATL) is widely used in the clinic, the mechanism underlying its high therapeutic efficacy against neuropathic pain remains unclear. NMDA receptors (NMDARs) represent a target for ATL and are involved in sensitization of neuropathic pain. Here we describe two actions of ATL on NMDARs: 1) enhancement of Ca(2+)-dependent desensitization and 2) trapping channel block. Inhibition of NMDARs by ATL was found to be dependent upon external Ca(2+) concentration ([Ca(2+)]) in a voltage-independent manner, with an IC(50) of 0.72 μM in 4 mM [Ca(2+)]. The ATL IC(50) value increased exponentially with decreasing [Ca(2+)], with an e-fold change observed per 0.69 mM decrease in [Ca(2+)]. Loading neurons with BAPTA abolished Ca(2+)-dependent inhibition, suggesting that Ca(2+) affects NMDARs from the cytosol. Since there is one known Ca(2+)-dependent process in gating of NMDARs, we conclude that ATL most likely promotes Ca(2+)-dependent desensitization. We also found ATL to be a trapping open-channel blocker of NMDARs with an IC(50) of 220 µM at 0 mV. An e-fold change in ATL IC(50) was observed to occur with a voltage shift of 50 mV in 0.25 mM [Ca(2+)]. Thus, we disclose here a robust dependence of ATL potency on extracellular [Ca(2+)], and demonstrate that ATL bound in the NMDAR pore can be trapped by closure of the channel.