Dataset Information

Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

ABSTRACT: Rodent genomic alignment sequences support a 2-exon model for muscarinic M4 receptor. Using this model a novel N-terminal extension was discovered in the human muscarinic acetylcholine M4 receptor. An open reading frame was discovered in the human, mouse and rat with a common ATG (methionine start codon) that extended the N-terminus of the muscarinic acetylcholine M4 receptor subtype by 155 amino acids resulting in a longer variant. Transcriptional evidence for this splice variant was confirmed by RNA-Seq and RT-PCR experiments performed from human donor brain prefrontal cortices. We detected a human upstream exon indicating the translation of the mature longer M4 receptor transcript. The predicted size for the longer two-exon M4 receptor splice variant with the additional 155 amino acid N-terminal extension, designated M4L is 69.7 kDa compared to the 53 kDa canonical single exon M4 receptor (M4S). Western blot analysis from a mammalian overexpression system, and saturation radioligand binding with [3H]-NMS (N-methyl-scopolamine) demonstrated the expression of this new splice variant. Comparative pharmacological characterization between the M4L and M4S receptors revealed that both the orthosteric and allosteric binding sites for both receptors were very similar despite the addition of an N-terminal extension.

SUBMITTER: Schober DA

PROVIDER: S-EPMC5718406 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

Schober Douglas A DA Croy Carrie H CH Ruble Cara L CL Tao Ran R Felder Christian C CC

PloS one 20171206 12

Rodent genomic alignment sequences support a 2-exon model for muscarinic M4 receptor. Using this model a novel N-terminal extension was discovered in the human muscarinic acetylcholine M4 receptor. An open reading frame was discovered in the human, mouse and rat with a common ATG (methionine start codon) that extended the N-terminus of the muscarinic acetylcholine M4 receptor subtype by 155 amino acids resulting in a longer variant. Transcriptional evidence for this splice variant was confirmed ...[more]

PMID: 29211764

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Project description:Accumulating evidence suggests that selective M4 muscarinic acetylcholine receptor (mAChR) activators may offer a novel strategy for the treatment of psychosis. However, previous efforts to develop selective M4 activators were unsuccessful because of the lack of M4 mAChR subtype specificity and off-target muscarinic adverse effects. We recently developed VU0152100, a highly selective M4 positive allosteric modulator (PAM) that exerts central effects after systemic administration. We now report that VU0152100 dose-dependently reverses amphetamine-induced hyperlocomotion in rats and wild-type mice, but not in M4 KO mice. VU0152100 also blocks amphetamine-induced disruption of the acquisition of contextual fear conditioning and prepulse inhibition of the acoustic startle reflex. These effects were observed at doses that do not produce catalepsy or peripheral adverse effects associated with non-selective mAChR agonists. To further understand the effects of selective potentiation of M4 on region-specific brain activation, VU0152100 alone and in combination with amphetamine were evaluated using pharmacologic magnetic resonance imaging (phMRI). Key neural substrates of M4-mediated modulation of the amphetamine response included the nucleus accumbens (NAS), caudate-putamen (CP), hippocampus, and medial thalamus. Functional connectivity analysis of phMRI data, specifically assessing correlations in activation between regions, revealed several brain networks involved in the M4 modulation of amphetamine-induced brain activation, including the NAS and retrosplenial cortex with motor cortex, hippocampus, and medial thalamus. Using in vivo microdialysis, we found that VU0152100 reversed amphetamine-induced increases in extracellular dopamine levels in NAS and CP. The present data are consistent with an antipsychotic drug-like profile of activity for VU0152100. Taken together, these data support the development of selective M4 PAMs as a new approach to the treatment of psychosis and cognitive impairments associated with psychiatric disorders such as schizophrenia.

Dataset Information

Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

Publications

Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

Similar Datasets

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