Reduced Chrna7 expression in mice is associated with decreases in hippocampal markers of inhibitory function: implications for neuropsychiatric diseases

Abstract

The α7* nicotinic acetylcholine receptor encoded by CHRNA7 (human)/Chrna7 (mice) regulates the release of both the inhibitory neurotransmitter GABA and the excitatory neurotransmitter glutamate in the hippocampal formation. A heterozygous (Het) deletion at 15q13.3 containing CHRNA7 is associated with increased risk for schizophrenia, autism, and epilepsy. Each of these diseases are characterized by abnormalities in excitatory and inhibitory hippocampal circuit function. Reduced Chrna7 expression results in decreased hippocampal α7* receptor density, abnormal hippocampal auditory sensory processing, and increased hippocampal CA3 pyramidal neuron activity in C3H mice Het for a null mutation in Chrna7. These abnormalities demonstrate that decreased Chrna7 expression alters hippocampal inhibitory circuit function. The current study examined the specific impact of reduced Chrna7 expression on hippocampal inhibitory circuits by measuring the levels of GABA, GABA(A) receptors, the GABA synthetic enzyme l-glutamic acid decarboxylase-65 (GAD-65), and the vesicular GABA transporter 1 (GAT-1) in wild-type (Chrna7 +/+) and Het (Chrna7 +/-) C3H α7 mice of both genders. GAD-65 levels were significantly decreased in male and female Het C3H α7 mice, whereas GABA(A) receptors were significantly reduced only in male Het C3H α7 mice. No changes in GABA and GAT-1 levels were detected. These data suggest that reduced CHRNA7 expression may contribute to the abnormalities in hippocampal inhibitory circuits observed in schizophrenia, autism, and/or epilepsy.

Figure 1

Photomicrographs of transverse sections through the hippocampus of adult C3H α7 mice labeled for glutamate decarboxylase-65 (GAD-65) (A1 – A4) or the GABA_A receptor antagonist [³⁵S]-tert-butylbicyclophosphorothionate ([³⁵S]-TBPS) (B1 – B4) A1. Transverse section through the hippocampal formation of a female wild type C3H α7 mouse immunolabeled for GAD-65. A2. Higher magnification of GAD-65 labeling in hippocampal area CA3 of the same section as in A1. A3. Transverse section through the hippocampal formation of a female heterozygous C3H α7 mouse immunolabeled for GAD-65. A4. Higher magnification of GAD-65 labeling in hippocampal area CA3 of the same section as in A3. Note the decrease in GAD-65 labeling in strata oriens, pyramidale, radiatum and lacunosum/moleculare of the heterozygous mouse (A3, A4) relative to the same regions in the wild type mouse (A1, A2). B1. Transverse section through the hippocampal formation of a male wild type C3H α7 mouse bound with the GABA_A antagonist [³⁵S]-TBPS. B2. Higher magnification of [³⁵S]-TBPS binding in the same section as in B1. B3. [³⁵S]-TBPS binding in a transverse section through the hippocampal formation of a male heterozygous C3H α7 mouse. B4. Higher magnification of [³⁵S]-TBPS binding in the same section as in B3. Note the decrease in [³⁵S]-TBPS binding in strata radiatum and lacunosum/moleculare of the heterozygous mouse (B3, B4) compared to the same regions in the wild type mouse (B1, B2). Abbreviations: GC - granule cell layer of the dentate gyrus, O - stratum oriens, P - stratum pyramidale, R - stratum radiatum, LM - stratum lacunosum/moleculare.

Figure 2

Photomicrographs of transverse sections through the hippocampus of adult C3H α7 mice labeled for γ-aminobutyric acid (GABA) (A1, A2) or the GABA vesicular transporter GAT-1 (B1, B2). A1. Transverse section through the hippocampal formation of a male wild type C3H α7 mouse labeled for GABA. A2. Transverse section through the hippocampal formation of a male heterozygous C3H α7 mouse labeled for GABA. The level of GABA labeling is comparable between the two genotypes. B1. Transverse section through the hippocampal formation of a male wild type C3H α7 mouse labeled for GAT-1. B2. Transverse section through the hippocampal formation of a male heterozygous C3H α7 mouse labeled for GAT-1. The level of GAT-1 labeling is comparable between the two genotypes. Abbreviations: GC - granule cell layer of the dentate gyrus, O - stratum oriens, P - stratum pyramidale, R - stratum radiatum, LM - stratum lacunosum/moleculare.

Figure 3

Qualitative measurement of immunostaining for the GABA synthesizing enzyme GAD-65 in hippocampal sections from female (n = 6/group, left panel) and male (n = 6/group, right panel) wild type and heterozygous C3H a7 mice. Decreased levels of GAD-65 immunostaining were seen in both male and female heterozygous mice, with significant (p < 0.05) decreases in strata lucidum/radiatum and lacunosum/moleculare of hippocampal area CA3 in mice of both genders. Abbreviations: DG MOL – molecular layer of the dentate gyrus, H – hilus (polymorph layer) of dentate gyrus, CA3 O – oriens layer of hippocampal area CA3, CA3 P – pyramidal cell layer of hippocampal area CA3, CA3 LR – lucidum/radiatum layer of hippocampal area CA3, CA3 LM – lacunosum moleculare layer of hippocampal area CA3, CA1 O – oriens layer of hippocampal area CA1, CA1 P – pyramidal cell layer of hippocampal area CA1, CA1 R – radiatum layer of hippocampal area CA1, CA1 LM – lacunosum moleculare of hippocampal area CA1. Values = mean ± S.E.M.

Figure 4

[35S]-TBPS binding in hippocampal sections from female (n = 6/group, left panel) and male (n = 6/group, right panel) wildtype and heterozygous C3H a7 mice. No significant differences in GABA_A receptor levels were seen in the female mice. In contrast, significant decreases in GABA_A receptor density were seen in the molecular layer of the dentate gyrus and in strata radiatum and lacunosum moleculare of hippocampal areas CA3 and CA1 in male heterozygous C3H a7 mice. Abbreviations: DG MOL – molecular layer of the dentate gyrus, H – hilus (polymorph layer) of dentate gyrus, CA3 O – oriens layer of hippocampal area CA3, CA3 R – radiatum layer of hippocampal area CA3, CA3 LM – lacunosum moleculare layer of hippocampal area CA3, CA1 O – oriens layer of hippocampal area CA1, CA1 R – radiatum layer of hippocampal area CA1, CA1 LM – lacunosum moleculare of hippocampal area CA1. Values = mean ± S.E.M.