Inhalational Anesthetics Induce Neuronal Protein Aggregation and Affect ER Trafficking

Matthew Coghlan,Christelle Petroz,Jason T Maynes,Sadiq Shaik,Pablo Rossi,Mark Crawford,Elizabeth Richards,Ramesh Babu Vanama,Saumel Ahmadi

doi:10.1038/s41598-018-23335-0

Abstract

Anesthetic agents have been implicated in the causation of neurological and cognitive deficits after surgery, the exacerbation of chronic neurodegenerative disease, and were recently reported to promote the onset of the neurologic respiratory disease Congenital Central Hypoventilation Syndrome (CCHS), related to misfolding of the transcription factor Phox2B. To study how anesthetic agents could affect neuronal function through alterations to protein folding, we created neuronal cell models emulating the graded disease severity of CCHS. We found that the gas anesthetic isoflurane and the opiate morphine potentiated aggregation and mislocalization of Phox2B variants, similar to that seen in CCHS, and observed transcript and protein level changes consistent with activation of the endoplasmic reticulum (ER) unfolded protein response. Attenuation of ER stress pathways did not result in a correction of Phox2B misfolding, indicating a primary effect of isoflurane on protein structure. We also observed that isoflurane hindered the folding and activity of proteins that rely heavily on ER function, like the CFTR channel. Our results show how anesthetic drugs can alter protein folding and induce ER stress, indicating a mechanism by which these agents may affect neuronal function after surgery.

Highlights

Extra alanines (33–35 total), a patient usually requires ventilatory support from birth, but a smaller number of alanines is associated with later disease onset, often occurring after a precipitating event with physiological stress[28,29]
Congenital Central Hypoventilation Syndrome (CCHS) disease severity is tied to the size of the alanine expansion present in the Phox2B transcription factor[27]
We observed that the smaller alanine constructs (20Ala, 23Ala, 25Ala, 27Ala and 30Ala) had 97–100% nuclear localization, consistent with the protein being a transcription factor, where the most severe 33Ala variant was present as cytosolic aggregates in approximately half of the cells (41.1% ± 0.9, mean ± s.e.m.)

Summary

Introduction

Extra alanines (33–35 total), a patient usually requires ventilatory support from birth, but a smaller number of alanines is associated with later disease onset, often occurring after a precipitating event with physiological stress[28,29]. The patient described to have anesthesia-induced onset of CCHS possessed five extra alanines (25 total), and was asymptomatic prior to anesthesia exposure[21]. Non-human disease models and post-mortem pathology reveals that CCHS is associated with Phox2B aggregation in the cytoplasm (as a transcription factor, it is usually nuclear), insinuating that anesthetic agents may be able to alter the folding/aggregation of the protein. This is interesting since many chronic neurodegenerative diseases, at least partially, result from protein misfolding, indicating that anesthesia effects on the folding and aggregation of proteins may be a contributive etiology to POCD. We found that modulation of ER stress cannot attenuate isoflurane-induced protein misfolding, illustrating a potential direct effect of the anesthetic drug on protein structure, with implications for mitigation or attenuation of POCD

Methods

Results

Conclusion