Actions of local anesthetics on NMDA receptor responses
Paganelli, Meaghan Ann
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Excitatory glutamatergic synaptic transmission plays a critical role in both proper central nervous system (CNS) development and function as well as neurological pathologies. In particular, the ionotropic glutamate receptor (iGluR) N -methyl-D-aspartate (NMDA) mediates the synaptic plasticity essential for learning and memory processes in the brain; however, this receptor also is responsible for the induction and maintenance of central sensitization, the underlying synaptic adaptations in the spinal cord dorsal horn that lead to the manifestation of chronic pain syndrome. Local anesthetics have been widely used in clinical practice to both prevent and manage acute and chronic pain. Canonically, local anesthetics inhibit the generation of action potentials by obstructing ion conduction through voltage-gated Na + channels, and, thus, impair signal transduction between neurons. Besides Na + channels, local anesthetics also act on a multiplicity of other ion channels, including the NMDA receptor. Recently, in the clinically relevant low mM range, studies have shown that amide-class local anesthetics bupivacaine and lidocaine inhibit NMDA-elicited currents. The inhibition is present in recombinant expression systems, pyramidal neurons and dorsal horn neurons in slices, suggesting that in physiological conditions NMDA receptors are affected by local anesthetics. However, the inhibitory mechanism employed by local anesthetics is poorly understood. To expound the intricacies of bupivacaine and lidocaine's inhibition of NMDA receptors, we investigated NMDA responses from the single-channel to whole-cell level in recombinantly expressed receptors in HEK 293 cells and examined the kinetic properties of inhibition. Both bupivacaine and lidocaine demonstrated voltage-dependence; acted through a blocking and allosteric effect; were state-independent, inhibiting both the closed and open receptor; and gained access to the channel through hydrophilic and hydrophobic pathways. Uniquely, while bupivacaine reduced channel open probability by a reduction in mean open time (MOT) and increase in mean closed time (MCT), lidocaine reduced channel open probability solely through a decrease in MOT. Bupivacaine penetrated more of the transmembrane electric field than lidocaine, suggesting that these two drugs have different sites of action. Mutation of two phenylalanine residues within the channel vestibule maintained sensitivity to lidocaine but weakened the channel's sensitivity to bupivacaine. This verifies that these drugs do not have the same binding site(s), which is distinct to the NMDA receptor as the local anesthetic binding site in Na + channel is overlapping. Overall, these studies provide insight into the actions of clinically-relevant local anesthetics during the alleviation of pain conditions. Elucidation of the distinctive mechanisms employed by local anesthetics of the same class provides valuable insight into chemical modification of molecules that could differentially alter NMDA receptor responses in synaptic environments induced by central sensitization. Providing a quantitative tool for predicting NMDA receptor-driven synaptic responses in the presence of local anesthetics, these studies help explicate how local anesthetics' dampening of NMDA receptor activity in the spinal cord can prevent, treat and even possibly reverse chronic pain.