Downregulation of calcium-dependent NMDA receptor desensitization by sodium-calcium exchangers: a role of membrane cholesterol

Abstract Background The plasma membrane Na+/Ca2+-exchanger (NCX) has recently been shown to regulate Ca2+-dependent N-methyl-d-aspartate receptor (NMDAR) desensitization, suggesting a tight interaction of NCXs and NMDARs in lipid nanoclasters or “rafts”. To evaluate possible role of this interaction we studied effects of Li+ on NMDA-elicited whole-cell currents and Ca2+ responses of rat cortical neurons in vitro before and after cholesterol extraction by methyl-β-cyclodextrin (MβCD). Results Substitution Li+ for Na+ in the external solution caused a concentration-dependent decrease of steady-state NMDAR currents from 440 ± 71 pA to 111 ± 29 pA in 140 mM Na+ and 140 mM Li+, respectively. The Li+ inhibition of NMDAR currents disappeared in the absence of Ca2+ in the external solution (Ca2+-free), suggesting that Li+ enhanced Ca2+-dependent NMDAR desensitization. Whereas the cholesterol extraction with MβCD induced a decrease of NMDAR currents to 136 ± 32 pA in 140 mM Na+ and 46 ± 15 pA in 140 mM Li+, the IC50 values for the Li+ inhibition were similar (about 44 mM Li+) before and after this procedure. In the Ca2+-free Na+ solution the steady-state NMDAR currents after the cholesterol extraction were 47 ± 6% of control values. Apparently this amplitude decrease was not Ca2+-dependent. In the Na+ solution containing 1 mM Ca2+ the Ca2+-dependent NMDAR desensitization was greater when cholesterol was extracted. Obviously, this procedure promoted its development. In agreement, Li+ and KB-R7943, an inhibitor of NCX, both considerably reduced NMDA-activated Ca2+ responses. The cholesterol extraction itself caused a decrease of NMDA-activated Ca2+ responses and, in addition, abolished the effects of Li+ and KB-R7943. The cholesterol loading into the plasma membrane caused a recovery of the KB-R7943 effects. Conclusions Taken together our data suggest that NCXs downregulate the Ca2+-dependent NMDAR desensitization. Most likely, this is determined by a tight functional interaction of NCX and NMDAR molecules because of their co-localization in membrane lipid rafts. The destruction of these rafts is accompanied by an enhancement of NMDAR desensitization and a loss of NCX-selective agent effects on NMDARs

Inhibition of Plasma Membrane Na/Ca-Exchanger by KB-R7943 or Lithium Reveals Its Role in Ca-Dependent N-methyl-D-aspartate Receptor Inactivation

ABSTRACT To evaluate the possible role of the plasma membrane N

Inhibition of Plasma Membrane Na/Ca-exchanger by KB-R7943 or Lithium Reveals its Role in Ca-dependent NMDAR Inactivation.* Running title: Mechanism of KB-R7943 and lithium effects on NMDARs

ABSTRAC

Mechanosensitive meningeal nociception via Piezo channels: Implications for pulsatile pain in migraine?

© 2019 Elsevier Ltd Background: Recent discovery of mechanosensitive Piezo receptors in trigeminal ganglia suggested the novel molecular candidate for generation of migraine pain. However, the contribution of Piezo channels in migraine pathology was not tested yet. Therefore, in this study, we explored a potential involvement of Piezo channels in peripheral trigeminal nociception implicated in generation of migraine pain. Methods: We used immunohistochemistry, calcium imaging, calcitonin gene related peptide (CGRP) release assay and electrophysiology in mouse and rat isolated trigeminal neurons and rat hemiskulls to study action of various stimulants of Piezo receptors on migraine-related peripheral nociception. Results: We found that essential (35%) fraction of isolated rat trigeminal neurons responded to chemical Piezo1 agonist Yoda1 and about a half of Yoda1 positive neurons responded to hypo-osmotic solution (HOS) and a quarter to mechanical stimulation by focused ultrasound (US). In ex vivo hemiskull preparation, Yoda1 and HOS largely activated persistent nociceptive firing in meningeal branches of trigeminal nerve. By using our novel cluster analysis of pain spikes, we demonstrated that 42% of fibers responded to Piezo1 agonist and 20% of trigeminal fibers were activated by Yoda1 and by capsaicin, suggesting expression of Piezo receptors in TRPV1 positive peptidergic nociceptive nerve fibers. Consistent with this, Yoda1 promoted the release of the key migraine mediator CGRP from hemiskull preparation. Conclusion: Taken together, our data suggest the involvement of mechanosensitive Piezo receptors, in particular, Piezo1 subtype in peripheral trigeminal nociception, which provides a new view on mechanotransduction in migraine pathology and suggests novel molecular targets for anti-migraine medicine

Mechanosensitive meningeal nociception via Piezo channels: Implications for pulsatile pain in migraine?

© 2019 Elsevier Ltd Background: Recent discovery of mechanosensitive Piezo receptors in trigeminal ganglia suggested the novel molecular candidate for generation of migraine pain. However, the contribution of Piezo channels in migraine pathology was not tested yet. Therefore, in this study, we explored a potential involvement of Piezo channels in peripheral trigeminal nociception implicated in generation of migraine pain. Methods: We used immunohistochemistry, calcium imaging, calcitonin gene related peptide (CGRP) release assay and electrophysiology in mouse and rat isolated trigeminal neurons and rat hemiskulls to study action of various stimulants of Piezo receptors on migraine-related peripheral nociception. Results: We found that essential (35%) fraction of isolated rat trigeminal neurons responded to chemical Piezo1 agonist Yoda1 and about a half of Yoda1 positive neurons responded to hypo-osmotic solution (HOS) and a quarter to mechanical stimulation by focused ultrasound (US). In ex vivo hemiskull preparation, Yoda1 and HOS largely activated persistent nociceptive firing in meningeal branches of trigeminal nerve. By using our novel cluster analysis of pain spikes, we demonstrated that 42% of fibers responded to Piezo1 agonist and 20% of trigeminal fibers were activated by Yoda1 and by capsaicin, suggesting expression of Piezo receptors in TRPV1 positive peptidergic nociceptive nerve fibers. Consistent with this, Yoda1 promoted the release of the key migraine mediator CGRP from hemiskull preparation. Conclusion: Taken together, our data suggest the involvement of mechanosensitive Piezo receptors, in particular, Piezo1 subtype in peripheral trigeminal nociception, which provides a new view on mechanotransduction in migraine pathology and suggests novel molecular targets for anti-migraine medicine