Abstract 15165: Nitric Oxide Synthase 1 Adaptor Protein is Involved in Neurogenic Qt Interval Prolongation via Glutamate-Mediated Neurotoxicity in the Nucleus Tractus Solitarius
Nitric oxide synthase 1 adaptor protein (NOS1AP) variants are strongly related to QT interval prolongation and sudden cardiac death (SCD) in humans. The mechanism is unknown. NOS1AP is tightly associated with NOS1, a neural form, highly expressed in the central nervous system. NOS1AP mRNA levels were examined using real time PCR, indicating that NOS1AP expression was 3-fold higher in brain nucleus tractus solitarius (NTS) than the heart in Sprague Dawley rats. NTS immunostaining indicated that NOS1AP is co-localized in neurons within the rat NTS. To study the cardiac regulatory action, lentiviral vectors containing NOS1AP-shRNA were constructed and microinjected into the NTS. Western blot revealed that NOS1AP levels were significantly decreased in the NTS by Lv-NOS1AP-shRNA injection. Blood pressure (BP), heart rate (HR), and ECG were recorded using radiotelemetry before and after NTS viral vector injection. No significant difference was observed in HR and BP between Lv-NOS1AP-shRNA injected and control rats. However, heart rate variability was significantly reduced and the QT interval was markedly increased in rats that received NTS microinjection of Lv-NOS1AP-shRNA. To study the underlying cellular mechanisms, we examined the effect of Lv-NOS1AP-shRNA on glutamate-induced neurotoxicity in neurons cultured from the rat NTS. Selective silencing of NOS1AP significantly increased glutamate-induced neurotoxicity. Immunoprecipitation results demonstrated that treatment of cells with Lv-NOS1AP-shRNA enhanced the association between the glutamate NMDA receptor and NOS1. This could be explained by molecular studies showing that NOS1AP and NOS1 have a similar docking domain, which allows them to bind comparatively to NMDA receptors. Our results also demonstrated that glutamate treatment increased intracellular calcium and nitric oxide levels; and that Lv-NOS1AP-shRNA treatment significantly elevated glutamate-induced NO production. In summary, knockdown of NOS1AP increases the association of NOS1 with NMDA receptors, causing overproduction of NO after stimulation with glutamate, leading to glutamate-induced neurotoxicity. Thus, altered NOS1AP in the NTS might play an important role in the pathogenesis of cardiac arrhythmia and SCD.
- © 2013 by American Heart Association, Inc.