A layer specific histological framework for synaptic quantification in hippocampal CA1.

Neural circuits mature through coordinated assembly of excitatory and inhibitory synapses, yet quantitative frameworks that preserve laminar and dorsoventral coordinates remain limited. Here, we establish a layer boundary-based pipeline for wide-field tiled immunohistochemistry to quantify PSD-95 and gephyrin puncta across stratum oriens, stratum radiatum, and stratum lacunosum-moleculare within mouse hippocampal CA1. In dorsal CA1, analysis across P8, P16, P24, and P80 revealed layer-dependent maturation patterns, with SLM exhibiting temporally inverted excitatory and inhibitory synaptic trajectories. Because prenatal valproic acid exposure is widely used to model autism-associated neurodevelopmental abnormalities and has been linked to hippocampal circuit disruption, we next examined whether synaptic organization is altered across CA1 layers and along the dorsoventral axis. At P56, a 2 × 2 design comparing control and prenatal VPA exposure across dorsal and ventral axes to assess axis dependent synaptic remodeling. Prenatal valproic acid exposure produced layer- and axis specific alterations in puncta density, mean area, and total area, suggesting nonuniform vulnerability of synaptic architecture. A conductance-based leaky integrate-and-fire model further provided a heuristic translation framework, highlighting membrane potential fluctuations as a candidate readout when excitatory and inhibitory components co-vary. These findings provides a practical foundation for integrating histological, functional, and developmental analyses of hippocampal CA1 circuits.