Promoter IV‐BDNF deficiency disturbs cholinergic gene expression of CHRNA5, CHRM2, and CHRM5: effects of drug and environmental treatments

Abstract

Brain-derived neurotrophic factor (BDNF) promotes maturation of cholinergic neurons. However, how activity-dependent BDNF expression affects specific cholinergic gene expression remains unclear. This study addressed this question by determining mRNA levels of 22 acetylcholine receptor subunits, the choline transporter (CHT), and the choline acetyltransferase (ChAT) in mice deficient in activity-dependent BDNF via promoter IV (KIV) and control wild-type mice. Quantitative RT-PCR revealed significant reductions in nicotinic acetylcholine receptor alpha 5 (CHRNA5) in the frontal cortex and hippocampus and M5 muscarinic acetylcholine receptor (CHRM5) in the hippocampus, but significant increases in M2 muscarinic acetylcholine receptor (CHRM2) in the frontal cortex of KIV mice compared to wild-type mice. Three-week treatments with fluoxetine, phenelzine, duloxetine, imipramine, or an enriched environment treatment (EET) did not affect the altered expression of these genes except that EET increased CHRNA5 levels only in KIV frontal cortex. EET also increased levels of CHRNA7, CHT, and ChAT, again only in the KIV frontal cortex. The imipramine treatment was most prominent among the four antidepressants; it upregulated hippocampal CHRM2 and frontal cortex CHRM5 in both genotypes, and frontal cortex CHRNA7 only in KIV mice. To our knowledge, this is the first evidence that BDNF deficiency disturbs expression of CHRNA5, CHRM2, and CHRM5. Our results suggest that promoter IV-BDNF deficiency – which occurs under chronic stress – causes cholinergic dysfunctions via these receptors. EET is effective on CHRNA5, while its compensatory induction of other cholinergic genes or drugs targeting CHRNA5, CHRM2, and CHRM5 may become an alternative strategy to reverse these BDNF-linked cholinergic dysfunctions.

BDNF deficiency and enriched environment treatment affect neurotransmitter gene expression differently across ages

“…In a pre‐test, we prescreened genes and omitted the genes of which expression levels were too low and unreliable (Ct values >27 after 14 cycle preamplification). As for acetylcholine genes, we selected six genes showing BDNF/EET effects in our previous study (Sakata & Overacre, 2017). Expression of total 81 genes of dopamine, serotonin, noradrenaline, glutamate, gamma aminobutyric acid (GABA), and acetylcholine and BDNF‐related systems were analyzed.…”

“…BDNF is critical for maturation and function of most types of neurons, including glutamatergic, GABAergic, serotonergic, dopaminergic, adrenergic, and cholinergic neurons (Alderson, Alterman, Barde, & Lindsay, 1990; Baker, Stanford, Brown, & Hagg, 2005; Ghosh, Carnahan, & Greenberg, 1994; Gottmann, Mittmann, & Lessmann, 2009; Guo, Hellard, Huang, & Katz, 2005; Hyman et al, 1991; Lyons et al, 1999; Mamounas et al, 2000; Marty, Berninger, Carroll, & Thoenen, 1996; Ward & Hagg, 2000). BDNF deficiency causes aberrant expression of genes related to these neurotransmitter systems (neurotransmitter genes) (Chourbaji et al, 2004; Djalali et al, 2005; Rios et al, 2006; Sakata & Duke, 2014; Sakata & Overacre, 2017; Tripp et al, 2012). EET can normalize the expression changes (Bredy, Zhang, Grant, Diorio, & Meaney, 2004; Komitova et al, 2013; Lee et al, 2013; Mehta‐Raghavan, Wert, Morley, Graf, & Redei, 2016; Rampon et al, 2000; Sakata & Overacre, 2017; Zhu et al, 2014).…”

“…BDNF deficiency causes aberrant expression of genes related to these neurotransmitter systems (neurotransmitter genes) (Chourbaji et al, 2004; Djalali et al, 2005; Rios et al, 2006; Sakata & Duke, 2014; Sakata & Overacre, 2017; Tripp et al, 2012). EET can normalize the expression changes (Bredy, Zhang, Grant, Diorio, & Meaney, 2004; Komitova et al, 2013; Lee et al, 2013; Mehta‐Raghavan, Wert, Morley, Graf, & Redei, 2016; Rampon et al, 2000; Sakata & Overacre, 2017; Zhu et al, 2014). However, these studies have been done mostly in young adult animals and the age dependency of the effects remains largely unknown.…”

The effect of enriched environment across ages: A study of anhedonia and BDNF gene induction

“…One microgram of total RNA was converted to cDNA (First Strand cDNA Synthesis Kit, Roche Applied Science, Indianapolis, Indiana). qRT‐PCR was performed using BioMark, as described previously . Briefly, 25 ng of cDNA was preamplified with pooled primers (200 nM) and PreAmp Master Mix (Applied Biosystems, Carlsbad, California) for 14 cycles of 95°C for 15 seconds and 60°C for 4 minutes.…”

“…qRT-PCR was performed using Bio-Mark, as described previously. [44][45][46] HGPRT were used for reference because these genes previously showed the least variation of expression in the hippocampus and frontal cortex among 6 tested housekeeping genes (cyclophilin D, Hypoxanthine-guanine phosphoribosyl-transferase (HGPRT), TATA sequence binding protein (TBP), β-actin, β-tubulin and S19). Relative gene expression values were determined by using the 2 −ΔΔCt method of Livak and Schmittgen.…”