Na⁺Channel Expression and Neuronal Function in the Na⁺/H⁺Exchanger 1 Null Mutant Mouse

Abstract

Mice lacking Na+/H+ exchanger 1 (NHE1) suffer from recurrent seizures and die early postnatally. Although the mechanisms for seizures are not well established, our previous electrophysiological work has shown that neuronal excitability and Na+ current density are increased in hippocampal CA1 neurons of these mutant mice. However, it is unknown whether this increased density is related to altered expression or functional regulation of Na+ channels. In this work, we asked three questions: is the increased excitability limited to CA1 neurons, is the increased Na+ current density related to an increased Na+ channel expression, and, if so, which Na+channel subtype(s) is upregulated? Using neurophysiological, autoradiographic, and immunoblotting techniques, we showed that both CA1 and cortical neurons have an increase in membrane excitability and Na+ current density; Na+ channel density is selectively upregulated in the hippocampus and cortex ( P < 0.05); and Na+ channel subtype I is significantly increased in the hippocampus and Na+channel subtype II is increased in the cortex. Our results demonstrate that mice lacking NHE1 upregulate their Na+ channel expression in the hippocampal and cortical regions selectively; this leads to an increase in Na+ current density and membrane excitability. We speculate that neuronal overexcitability due to Na+ channel upregulation in the hippocampus and cortex forms the basis of epileptic seizures in NHE1 mutant mice.

Regulation of Early Neurite Morphogenesis by the Na+/H+ Exchanger NHE1

“…Similarly, although NHE1 is required for maximally efficient cell migration in non-neuronal cells, it is not essential for the development of the migratory response (Hayashi et al, 2008). The fact that NHE1-null mice, although exhibiting a distinctive neurological phenotype that includes ataxia, truncal instability, seizures, and selective neuronal death, do not display gross neurodevelopmental defects (Cox et al, 1997;Bell et al, 1999) also points to a permissive role for NHE1 in neurite morphogenesis, although it must be noted that interpretation of the NHE1 Ϫ/Ϫ phenotype is complicated by alterations in the expression and/or activities of other pH i regulating mechanisms and Na ϩ influx pathways in NHE1 Ϫ/Ϫ neurons that compensate for the loss of NHE1 (Xia et al, 2003;Xue et al, 2003;Schwab et al, 2005) (see also Putney and Barber, 2004;Zhou et al, 2004). Similarly, mice lacking the Cl Ϫ /HCO 3 Ϫ exchanger AE3 (Hentschke et al, 2006), the Na ϩ -K ϩ -2Cl Ϫ cotransporter NKCC1 (Flagella et al, 1999), or the K ϩ -Cl Ϫ cotransporter KCC2 (Hübner et al, 2001) do not exhibit gross histological abnormalities, despite the fact that these transport mechanisms are also known to regulate neurite outgrowth (Nakajima et al, 2007;Pieraut et al, 2007;Blaesse et al, 2009).…”

“…Neocortical neurons were isolated from either embryonic day 16 (E16) wild-type (WT) C57BL/6 mice (Charles River Laboratories) or postnatal day 0.5 (P0.5) Nhe1 homozygous mutant (NHE1 Ϫ/Ϫ ), heterozygous (NHE1 ϩ/Ϫ ), and wild-type (NHE1 ϩ/ϩ ) littermate progeny of matings between heterozygous B6.SJL, ϩ/swe mice (The Jackson Laboratory), which harbor a spontaneous mutation in the Nhe1 allele (Cox et al, 1997). In the latter cases, animals were killed within 12 h of birth, and genotyping was performed on brain tissue as described previously (Xue et al, 2003) using the following primers: sense, 5Ј-CACTCTCTGCATCCCTCCTC-3Ј and antisense, 5Ј-AAGTCAT-GCGGCAAGCTAGT-3Ј, corresponding to base pairs 47312-47331 and 47956-47977 of the intronic region of the mouse NHE1 cDNA sequence (GenBank accession number BC052708). To isolate neocortical neurons, cortices freed of meninges were placed in ice-cold (4°C) HBSS supplemented with 10% FBS and penicillin/streptomycin and gently triturated with fire-polished Pasteur pipettes of diminishing tip diameter.…”

Decreased Neuronal Death in Na+/H+Exchanger Isoform 1-Null Mice afterIn VitroandIn VivoIschemia

“…The combination of these approaches overcame the shortcomings of each method. For instance, the spontaneous NHE1 mutant mice (slow-wave epilepsy) exhibit upregulation of NHE3 in cerebellum, Na ϩ channel current density in hippocampal and cortical neurons, and downregulation of brain-specific anion exchanger isoform 3 in the hippocampus (Xia et al, 2003;Xue et al, 2003). In addition, NHE1-deficient brains exhibited regionally specific downregulation of many genes, implying that NHE1 may function as a signaling molecule (Zhou et al, 2004).…”

Traditional and emerging roles for the SLC9 Na+/H+ exchangers

“…This phenotpye was associated with selective neuronal death in the cerebellum and brainstem of KO mice [34]. KO of NHE1 decreased steady-state pH i , attenuated pH i recovery from cell acidification (even in the presence of HCO 3 -) and increased expression and current density of voltage-gated Na + channels in hippocampal and cortical regions [177,56,185]. Thus, loss of NHE1 seems to alter expression and activity of other membrane transport proteins in the brain, resulting in increased neuronal excitability.…”