Anti-peptide antibodies that specifically recognize the alpha1 subunit of class A-D voltage-gated Ca2+ channels and a monoclonal antibody (MANC-1) to the alpha2 subunit of L-type Ca2+ channels were used to investigate the distribution of these Ca2+ channel subtypes in neurons and glia in models of brain injury, including kainic acid-induced epilepsy in the hippocampus, mechanical and thermal lesions in the forebrain, hypomyelination in white matter, and ischemia. Immunostaining of the alpha2 subunit of L-type Ca2+ channels by the MANC-1 antibody was increased in reactive astrocytes in each of these forms of brain injury. The alpha1C subunits of class C L-type Ca2+ channels were upregulated in reactive astrocytes located in the affected regions in each of these models of brain injury, although staining for the alpha1 subunits of class D L-type, class A P/Q-type, and class B N-type Ca2+ channels did not change from patterns normally observed in control animals. In all of these models of brain injury, there was no apparent redistribution or upregulation of the voltage-gated Ca2+ channels in neurons. The upregulation of L-type Ca2+ channels in reactive astrocytes may contribute to the maintenance of ionic homeostasis in injured brain regions, enhance the release of neurotrophic agents to promote neuronal survival and differentiation, and/or enhance signaling in astrocytic networks in response to injury.

The Emx1-IRES-Cre transgenic mouse is commonly used to direct genetic recombination in forebrain excitatory neurons. However the original study reported that Emx1-Cre is also expressed in peripheral autonomic ganglia, which could potentially affect the interpretation of targeted circuitry contributing to systemic phenotypes. Here, we report that Emx1-Cre is expressed in nodose ganglion cells whose axon travel within afferent vagal nerves and terminate in the nucleus tractus solitarius (NTS), a critical central autonomic regulatory pathway located in the dorsal medulla. Optogenetic stimulation of Emx1-Cre+ fibers reliably evoked excitatory postsynaptic currents in the NTS in vitro and evoked a parasympathetic bradycardia reflex occasionally with irregular respirations in anesthetized mice in vivo. Our study clearly demonstrates that Emx1-IRES-Cre is expressed in the vagal nerve fibers traveling within the central autonomic system involving autonomic cardiorespiratory regulation. These results raise caution when interpreting the systemic phenotypes of Emx1-IRES-Cre conditional recombinant mice, and also suggest the utility of this line to investigate the modulators of afferent vagal system.Significance StatementEmx1-IRES-Cre mice are widely used to dissect critical circuitry underlying neurological disorders such as epilepsy. These studies often assume the Cre is expressed selectively only in forebrain excitatory neurons. However, earlier works reported that Emx1 is expressed in several peripheral tissues of the developing embryo and thus gene recombination may affect these peripheral structures. In this study, we characterized the expression and physiological functions of Emx1-Cre expressed in the afferent vagus nerve. Optogenetic stimulation of these Emx1-Cre+ vagus nerves activates the nucleus tractus solitarius neurons within the brainstem in vitro and induces brief bradycardia in vivo. This study confirmed that peripheral Emx1-Cre+ cells are involved in autonomic regulation and potentially affect transgenic mouse phenotypes.

ANK3 is a leading bipolar disorder candidate gene in humans. Previous studies showed that deletion of Ank3-1b, a bipolar disorder (BD)-associated variant of Ank3 in mice leads to increased firing threshold and diminished action potential dynamic range of parvalbumin (PV)-interneurons and absence epilepsy, thus providing a biological mechanism linking epilepsy and BD. To better understand the impact of defective PV interneurons on network activity and behavior in these mice, we examined spectral EEG correlates of behaviors seen in Ank3-1b knockout (KO) mice during home-cage activity using paired video-EEG recordings. Since PV-interneurons contribute to the generation of high frequency gamma oscillations, we anticipated changes in the power of EEG signals in the gamma frequency range (> 25 Hz) during behavioral deficits related to human BD symptoms, including abnormal sleep and activity levels. Ank3-1b KO mice exhibited an overall increase in slow gamma (~25-45 Hz) power compared to controls, and slow gamma power correlated with seizure phenotype severity across behaviors. During sleep, increased slow gamma power correlated with decreases in time spent in the rapid eye movement (REM) stage of sleep. Seizures were more common during REM sleep compared to non-REM (NREM) sleep. We also found that Ank3-1b KO mice were hyperactive and exhibited a repetitive behavior phenotype that correlated with increased slow gamma power. Our results identify a novel EEG biomarker associating Ank3 genetic variation with BD and suggest modulation of gamma oscillations as a potential therapeutic target.