Neurochemistry of the afferents to the rat cochlear root nucleus: Possible synaptic modulation of the acoustic startle

Abstract

Afferents to the primary startle circuit are essential for the elicitation and modulation of the acoustic startle reflex (ASR). In the rat, cochlear root neurons (CRNs) comprise the first component of the acoustic startle circuit and play a crucial role in mediating the ASR. Nevertheless, the neurochemical pattern of their afferents remains unclear. To determine the distribution of excitatory and inhibitory inputs, we used confocal microscopy to analyze the immunostaining for vesicular glutamate and GABA transporter proteins (VGLUT1 and VGAT) on retrogradely labeled CRNs. We also used reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry to detect and localize specific neurotransmitter receptor subunits in the cochlear root. Our results show differential distributions of VGLUT1- and VGAT-immunoreactive endings around cell bodies and dendrites. The RT-PCR data showed a positive band for several ionotropic glutamate receptor subunits, M1-M5 muscarinic receptor subtypes, the glycine receptor alpha1 subunit (GlyRalpha1), GABAA, GABAB, and subunits of alpha2 and beta-noradrenergic receptors. By immunohistochemistry, we confirmed that CRN cell bodies exhibit positive immunoreaction for the glutamate receptor (GluR) 3 and NR1 GluR subunits. Cell bodies and dendrites were also positive for M2 and M4, and GlyRalpha1. Other subunits, such as GluR1 and GluR4 of the AMPA GluRs, were observed in glial cells neighboring unlabeled CRN cell bodies. We further confirmed the existence of noradrenergic afferents onto CRNs from the locus coeruleus by combining tyrosine hydroxylase immunohistochemistry and tract-tracing experiments. Our results provide valuable information toward understanding how CRNs might integrate excitatory and inhibitory inputs, and hence how they could elicit and modulate the ASR.

Origin and function of short-latency inputs to the neural substrates underlying the acoustic startle reflex

“…As expected from our previous report (Gómez-Nieto et al, 2008b), we found a strong immunoreactivity for VGLUT1 in the cochlear root nucleus (Supplemental Figure 1). VGLUT1-labeled endings equally distributed throughout dorsal and ventral portions of the cochlear root nucleus (Supplemental Figure 1).…”

Auditory System

“…The cochlear root neurons show a latency of 2-3 ms and they respond with a "primary-like with notch" pattern to pure tone stimulation (Sinex et al, 2001). These data suggest that they participate in the acoustic startle reflex Friauf, 1992, 1994;Lee et al, 1996;Gómez-Nieto et al, 2008a, 2008b.…”

“…This is the so-called acoustic nerve nucleus or interstitial nucleus of the vestibulocochlear nerve (mouse: Lorente de Nó, 1926;rat: Harrison and Warr, 1962;Harrison and Feldman, 1970;Merchán et al, 1988;Osen et al, 1991;López et al, 1999). These neurons receive axon collaterals terminating in small boutons arising from the main stem axon of the primary afferents and other noradrenergic and cholinergic afferents from the locus coeruleus and ventral nucleus of the trapezoid body, respectively (López et al, 1999, Gómez-Nieto et al, 2008a, 2008b (Fig. 5).…”

Ultrastructure, synaptic organization, and molecular components of bushy cell networks in the anteroventral cochlear nucleus of the rhesus monkey

“…Free-floating brainstem sections followed two cycles of freezing and thawing on dry ice, were washed ipn 0.1 M PB and incubated with the corresponding primary antibody (Table 1) as described previously (Gómez-Nieto et al 2008b; Rubio et al, 2008; Gómez-Nieto and Rubio, 2009). Briefly, all immunostaining steps were performed at room temperature (~22°C) unless otherwise stated.…”