Abstract
INTRODUCTION: The rostral ventrolateral medulla (RVLM) is a vital vasomotor nucleus involved in hypertension control. However, the molecular intricacies within RVLM cells, particularly in non-neuronal cell populations such as astrocytes, under hypertensive conditions remain to be systematically investigated. OBJECTIVES: We aimed to elucidate cellular heterogeneity in the RVLM at single-cell resolution, with a focus on astrocytes, to further characterize its role in hypertension. METHODS: Transcriptional changes in cells in the RVLM after hypertension were examined by using single-nucleus RNA sequencing (snRNA-seq). Further experiments, like radiotelemetry and immunofluorescence assays, were conducted to support the findings of snRNA-seq. RESULTS: In total, 36 982 high-quality nuclei were obtained, with identification of nine cell types. Three transcriptomically distinct subtypes of astrocytes were defined within the astrocyte cell population: Cntnap2(+), Pld5(+), and Ppp1r16b(+). The expression of Cntnap2 in RVLM astrocytes was notably increased to approximately 2.0-fold (P < 0.001) upon hypertension. Astrocyte-specific Cntnap2 overexpression in RVLM of WKY rats significantly enhanced neuronal excitability (∼21.5-fold, P < 0.001), sympathetic outflow (∼4.2-fold, P < 0.001), and blood pressure (BP, ∼1.5-fold, P < 0.001). Astrocyte-specific Cntnap2 knockdown in RVLM of SHRs resulted in 71 % reduction in neuronal excitability (P < 0.001), 51.6 % decrease in sympathetic outflow (P < 0.01), and 12.6 % drop in BP (P < 0.01). Elevated levels of Cntnap2 in primary astrocytes markedly increased primary neuron excitability to approximately 8.2-fold (P < 0.001). Mechanistically, Cntnap2 upregulation decreased Eaat2 expression by 26.5 % (P < 0.001), impairing astrocytic glutamate uptake. Importantly, reintroducing Eaat2 into astrocytes blocked 90.5 % (P < 0.001) of Cntnap2-induced neuronal excitability. CONCLUSION: Our findings suggest that Cntnap2 may impede glutamate homeostasis at least partially through regulating Eaat2 in astrocytes, which could consequently enhance RVLM neuronal excitability, increase sympathetic tone, and contribute to elevated BP.