Differential expressions of the alternatively spliced variant mRNAs of the µ opioid receptor gene, OPRM1, in brain regions of four inbred mouse strains.

Gavril W Pasternak,Benjamin Kest,Amanda R Waxman,Zhigang Lu,Grace C Rossi,Jin Xu,Mingming Xu,Tuck Wah Soong,Ying-Xian Pan

doi:10.1371/journal.pone.0111267

Gavril W Pasternak, Benjamin Kest + Show 7 more

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https://doi.org/10.1371/journal.pone.0111267

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Abstract

The µ opioid receptor gene, OPRM1, undergoes extensive alternative pre-mRNA splicing in rodents and humans, with dozens of alternatively spliced variants of the OPRM1 gene. The present studies establish a SYBR green quantitative PCR (qPCR) assay to more accurately quantify mouse OPRM1 splice variant mRNAs. Using these qPCR assays, we examined the expression of OPRM1 splice variant mRNAs in selected brain regions of four inbred mouse strains displaying differences in µ opioid-induced tolerance and physical dependence: C56BL/6J, 129P3/J, SJL/J and SWR/J. The complete mRNA expression profiles of the OPRM1 splice variants reveal marked differences of the variant mRNA expression among the brain regions in each mouse strain, suggesting region-specific alternative splicing of the OPRM1 gene. The expression of many variants was also strain-specific, implying a genetic influence on OPRM1 alternative splicing. The expression levels of a number of the variant mRNAs in certain brain regions appear to correlate with strain sensitivities to morphine analgesia, tolerance and physical dependence in four mouse strains.

Highlights

Morphine and most clinical analgesic agents, as well as heroin, act primarily through m opioid receptors [1]
These observations are consistent with the concept of subtypes of m opioid receptors that was initially proposed by pharmacological studies [10,11,12] and subsequently confirmed with the isolation of a multitude of m opioid receptor variants generated by alternative splicing [13,14,15,16,17] in rodents and humans despite the presence of a single m opioid receptor gene, as determined by chromosomal mapping [18,19,20,21] and genomic sequencing
Using the established RT-SYBR green quantitative PCR (qPCR) assays, we examined the expression levels of OPRM1 splice variant mRNAs in nine brain regions of four inbred mouse strains

Summary

Introduction

Morphine and most clinical analgesic agents, as well as heroin, act primarily through m opioid receptors [1] Their actions, such as analgesia, tolerance, physical dependence and other side-effects vary among patients [2,3,4]. Studies show that the mouse, rat and human OPRM1 genes generate 29, 16 and 19 splice variants, respectively, with similar patterns (Fig. 1) (see reviews: [16,22] ) These splice variants can be categorized into three major groups based upon receptor structure: 1) Fulllength carboxyl (C-) terminal variants with 7-transmembrane (7TM) domains; 2) Truncated variants containing 6-TM domains; and 3) Truncated variants containing a single TM. The significance of the exon 11-associated 6-TM variants is illustrated by their role in the analgesic actions of opioids such as morphine-6b-glucuronide (M6G), fentanyl and heroin [33], as a novel new class of opioid analgesics lacking many of the side-effects of traditional opiates [34]

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