Deciphering the role of brainstem glycinergic neurons during startle and prepulse inhibition

Prepulse inhibition (PPI) of the auditory startle response is the gold standard operational measure of sensorimotor gating. Affected by various neurological and neuropsychiatric illnesses, PPI also declines during aging. While PPI deficits are often associated with cognitive overload, attention impairments and motor dysfunctions, their reversal is routinely used in experimental systems for drug screening. Yet, the cellular and circuit-level mechanisms of PPI remain unclear, even under non-pathological conditions. Recent evidence shows that neurons located in the brainstem caudal pontine reticular nucleus (PnC) expressing the glycine transporter type 2 (GlyT2 ⁺ ) receive inputs from the central nucleus of the amygdala (CeA) and contribute to PPI via an uncharted pathway. Using tract-tracing and immunohistochemical analyses in GlyT2-eGFP mice, we reveal the neuroanatomical location of CeA glutamatergic neurons innervating GlyT2 ⁺ neurons. Our precise in vitro optogenetic manipulations coupled to field electrophysiological recordings demonstrate that CeA glutamatergic inputs do suppress auditory neurotransmission in PnC neurons but not via action on transmitter release from auditory afferents. Rather, our data is consistent with excitatory drive onto GlyT2 ⁺ neurons. Indeed, our PPI experiments in vivo demonstrate that optogenetic activation of GlyT2 ⁺ PnC neurons increases PPI and is sufficient to induce PPI, clarifying the crucial role of these neurons in young GlyT2-Cre mice. In contrast, in older GlyT2-Cre mice, PPI is reduced and not further altered by optogenetic inhibition of GlyT2 ⁺ neurons. We conclude that GlyT2 ⁺ PnC neurons innervated by CeA glutamatergic inputs are crucial for PPI and we highlight their reduced activity during the age-dependent decline in PPI.