Background Prader-Willi syndrome (PWS) is a neurodevelopmental disorder caused by loss of paternally expressed genes on chromosome 15q11–13, including NDN, which encodes necdin. Necdin deficiency has been linked to impaired visuospatial memory, social recognition, and stress regulation—features also seen in PWS. Previous work showed that necdin-deficient (Ndn + m/−p) mice exhibit reduced activity of noradrenergic neurons in the locus coeruleus (LC), a nucleus essential for arousal and stress responses. However, the mechanisms underlying LC hypoactivity remain unclear. Because GABAergic signaling is critical for LC excitability, this study examined the role of GABAA and GABAB receptor-mediated inhibition in Ndn + m/−p mice. Methods Electrophysiological recordings from brainstem slices of wild-type (WT) and Ndn + m/−p mice were used to measure spontaneous firing rates (SFRs) of LC noradrenergic neurons. The effects of bicuculline (GABAA antagonist) and CGP54626 (GABAB antagonist) were tested. Whole-cell patch-clamp recordings assessed receptor-mediated currents. Western blotting quantified receptor subunit expression in peri-LC tissue. Immunocytochemistry and ELISA examined GABAB receptor expression and GABA release in cultured astrocytes. Results LC-NE neurons in Ndn + m/−p mice exhibited significantly reduced baseline SFR compared with WT. Bicuculline did not alter firing in either genotype, whereas CGP54626 significantly increased SFR in WT but not in Ndn + m/−p neurons, indicating impaired GABAB receptor-mediated tonic inhibition. Whole-cell patch-clamp experiments confirmed intact GABAA receptor-mediated inward currents in both genotypes, while no GABAB receptor-mediated phasic currents were detected. Western blot analysis revealed comparable expression of GABAA receptor α2 subunit and GABABR1 in peri-LC tissue between WT and Ndn + m/−p mice, suggesting functional rather than expression-level deficits. GFAP-positive cell density in the LC region was unchanged in vivo; however, in astrocyte cultures, Ndn + m/−p astrocytes exhibited greater proliferation by DIV 19 and consistently secreted higher levels of GABA, with significant elevations at later culture stages. Conclusion Necdin deficiency selectively disrupts GABAB receptor-mediated tonic inhibition of LC-NE neurons while preserving GABAA receptor function. Elevated astrocytic proliferation and GABA release may further enhance ambient inhibition, contributing to LC hypoactivity and the neurobehavioral phenotypes of PWS. These findings identify GABAB receptor dysfunction and astrocytic dysregulation as potential mechanistic targets for therapeutic intervention in PWS.

GABAergic regulation of Locus coeruleus activity in necdin-deficient mice, an animal model of Prader-Willi syndrome

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2025年12月30日

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Li-Ping Tsai,
Hao Chan,
Wei-Chen Hung,
Ming-Yuan Min,
Sin-Jhong Cheng,
Chen-En Yang,
Chun-Hsien Yu,
Shi-Bing Wong

摘要

Background

Prader-Willi syndrome (PWS) is a neurodevelopmental disorder caused by loss of paternally expressed genes on chromosome 15q11–13, including NDN, which encodes necdin. Necdin deficiency has been linked to impaired visuospatial memory, social recognition, and stress regulation—features also seen in PWS. Previous work showed that necdin-deficient (Ndn + m/−p) mice exhibit reduced activity of noradrenergic neurons in the locus coeruleus (LC), a nucleus essential for arousal and stress responses. However, the mechanisms underlying LC hypoactivity remain unclear. Because GABAergic signaling is critical for LC excitability, this study examined the role of GABA_A and GABA_B receptor-mediated inhibition in Ndn + m/−p mice.

Methods

Electrophysiological recordings from brainstem slices of wild-type (WT) and Ndn + m/−p mice were used to measure spontaneous firing rates (SFRs) of LC noradrenergic neurons. The effects of bicuculline (GABA_A antagonist) and CGP54626 (GABA_B antagonist) were tested. Whole-cell patch-clamp recordings assessed receptor-mediated currents. Western blotting quantified receptor subunit expression in peri-LC tissue. Immunocytochemistry and ELISA examined GABA_B receptor expression and GABA release in cultured astrocytes.

Results

LC-NE neurons in Ndn + m/−p mice exhibited significantly reduced baseline SFR compared with WT. Bicuculline did not alter firing in either genotype, whereas CGP54626 significantly increased SFR in WT but not in Ndn + m/−p neurons, indicating impaired GABA_B receptor-mediated tonic inhibition. Whole-cell patch-clamp experiments confirmed intact GABA_A receptor-mediated inward currents in both genotypes, while no GABA_B receptor-mediated phasic currents were detected. Western blot analysis revealed comparable expression of GABA_A receptor α2 subunit and GABA_BR1 in peri-LC tissue between WT and Ndn + m/−p mice, suggesting functional rather than expression-level deficits. GFAP-positive cell density in the LC region was unchanged in vivo; however, in astrocyte cultures, Ndn + m/−p astrocytes exhibited greater proliferation by DIV 19 and consistently secreted higher levels of GABA, with significant elevations at later culture stages.

Conclusion

Necdin deficiency selectively disrupts GABA_B receptor-mediated tonic inhibition of LC-NE neurons while preserving GABA_A receptor function. Elevated astrocytic proliferation and GABA release may further enhance ambient inhibition, contributing to LC hypoactivity and the neurobehavioral phenotypes of PWS. These findings identify GABA_B receptor dysfunction and astrocytic dysregulation as potential mechanistic targets for therapeutic intervention in PWS.