Expression of acid-sensing ion channels and selection of reference genes in mouse and naked mole rat

Abstract

Acid-sensing ion channels (ASICs) are a family of ion channels comprised of six subunits encoded by four genes and they are expressed throughout the peripheral and central nervous systems. ASICs have been implicated in a wide range of physiological and pathophysiological processes: pain, breathing, synaptic plasticity and excitotoxicity. Unlike mice and humans, naked mole-rats do not perceive acid as a noxious stimulus, even though their sensory neurons express functional ASICs, likely an adaptation to living in a hypercapnic subterranean environment. Previous studies of ASIC expression in the mammalian nervous system have often not examined all subunits, or have failed to adequately quantify expression between tissues; to date there has been no attempt to determine ASIC expression in the central nervous system of the naked mole-rat. Here we perform a geNorm study to identify reliable housekeeping genes in both mouse and naked mole-rat and then use quantitative real-time PCR to estimate the relative amounts of ASIC transcripts in different tissues of both species. We identify RPL13A (ribosomal protein L13A) and CANX (calnexin), and β-ACTIN and EIF4A (eukaryotic initiation factor 4a) as being the most stably expressed housekeeping genes in mouse and naked mole-rat, respectively. In both species, ASIC3 was most highly expressed in dorsal root ganglia (DRG), and ASIC1a, ASIC2b and ASIC3 were more highly expressed across all brain regions compared to the other subunits. We also show that ASIC4, a proton-insensitive subunit of relatively unknown function, was highly expressed in all mouse tissues apart from DRG and hippocampus, but was by contrast the lowliest expressed ASIC in all naked mole-rat tissues.

Acid and inflammatory sensitisation of naked mole-rat colonic afferent nerves

“…Recent single-cell RNA-sequencing analysis of colonic sensory neurones shows discrete expression of such acid-sensitive ion channels with differing populations of mouse colonic afferents, suggesting functional specialism [16]. Compared to mice, NMRs display a similar expression profile of ASICs throughout the nervous system [17], and of those analysed, with the exception of ASIC3, NMR acid sensors show similar activation profiles to those of mice [13,18,19]. NMR TRPV1 is also expressed in sensory afferents and shows similar proton sensitivity to mouse TRPV1 [18].…”

Naked mole-rat acid-sensing ion channel 3 forms nonfunctional homomers, but functional heteromers

“…We have previously demonstrated that naked mole-rat dorsal root ganglion (DRG) neurons produce ASIC-like currents in response to acid (36) and that these neurons express nmrASIC3 mRNA (33,36) and thus we used APETx2, an inhibitor of most ASIC3-containing ASIC trimers, to determine if nmrASIC3 contributes to the acidsensitivity of these neurons. A pH 5.0 stimulus evoked two types of inward current in naked molerat DRG neurons: rapidly-inactivating, ASIC-like currents and sustained, TRPV1-like currents ( Fig.…”

“…Understanding the structure-function of ASIC3 is of particular interest because there is substantial evidence supporting involvement of ASIC3 in pain (9,(23)(24)(25)(26)(27)(28), as well itch (29), mechanosensation (23,30) and anxiety (31). Although ASIC3 expression was initially thought to be restricted to the peripheral nervous system (and hence its original name, dorsal root ganglia acid-sensing ion channel, DRASIC (32)), we and others have demonstrated that ASIC3 is also expressed in both the spinal cord and numerous brain regions (33,34), which makes it even more important to understand how ASIC3 functions because any potential drug that targets ASIC3 for the treatment of pain may also produce side effects with the central nervous system.…”

“…Although naked mole-rats can more efficiently buffer CO 2 , our previous data regarding the inability of acid to evoke action potentials in an ex vivo preparation (35) that arises from an amino acid variation in NaV1.7 (36) demonstrates that there are likely multiple adaptations to living in a hypercapnic environment. We recently compared ASIC expression between mice and naked mole-rats, finding that whereas ASIC4 is highly abundant in mouse tissues it is the most lowly expressed ASIC transcript in naked mole-rat tissues; the expression pattern of ASIC3 was similar between species (33). At a functional level, we have previously shown that naked mole-rat ASIC1a, ASIC1b and TRPV1 are largely indistinguishable from the mouse orthologs (36,41) and here we set out to explore the function of naked mole-rat ASIC3 considering its importance to a physiological and pathophysiological processes.…”

Inhibition of Brain Asics Affects Hippocampal Theta-Rhythm and Openfield Behavior in Rats

“…Як було зазначено вище, ASIC-канали залучені у регуляції багатьох фізіологічних та патофізіологічних процесів, що відбуваються у ЦНС, а отже їхнє пригнічення ймовірно впливатиме на активність певних структур мозку, що в свою чергу, може супроводжуватися певними змінами у поведінці. Відомо, що гіпокамп бере участь у процесах навчання, запам'ятовування та реагування на новизну, а його нейрони характеризуються високим рівнем експресії ASIC1a-каналів [11].…”