Electrophysiological characterization of neuropeptide S-expressing neurons in the periLC: influence of the female estrous cycle on neuronal excitability in mice

Abstract Neuropeptide S (NPS) signaling is critically involved in arousal, sleep–wake regulation, fear and anxiety, and related forms of learning and memory, yet comparatively little is known about whether intrinsic properties of identified NPS neurons differ between sexes or fluctuate across the female estrous cycle. Here, we performed whole-cell patch-clamp recordings from genetically identified NPS-eGFP neurons in acute horizontal brainstem slices containing the peri–locus coeruleus (periLC) region. We show that periLC NPS neurons can be subdivided into two electrophysiologically distinct classes based on their discharge patterns from hyperpolarized membrane potentials: a delayed-onset firing phenotype (type I) and a regular-spiking phenotype (type II). Pharmacological and voltage-clamp analyses indicate that differential contribution of a transient A-type potassium current (I A ), consistent with Kv4 channel function, is a key determinant of these firing phenotypes and provides a mechanism for gain control in type I neurons. Across male and female mice, we did not detect robust sex differences in the relative abundance of these neuronal classes or in overall excitability measures, suggesting that baseline intrinsic properties of periLC NPS neurons are broadly comparable between sexes. In contrast, estrous staging revealed a selective modulation of intrinsic discharge in females: during non-receptive stages (metestrus/diestrus), type I neurons exhibited higher spike output and increased instantaneous firing frequencies compared with receptive stages (proestrus/estrus), whereas type II neurons were comparatively stable across the cycle. These findings provide a cellular mechanism by which ovarian state may tune NPS neuron output and, consequently, the impact of NPS signaling on downstream circuits controlling arousal and stress-related behaviors.