Cholinergic input from the ventral nucleus of the trapezoid body to cochlear root neurons in rats

Abstract

Brain stem pathways are essential for the modulation of the acoustic startle reflex by sounds; nevertheless, the neural circuits that convey fast auditory information to the primary acoustic startle circuit are still unclear. In the rat, cochlear root neurons (CRNs) comprise the first component of the primary acoustic startle circuit and are critical in the initiation and full expression of the acoustic startle reflex. To determine whether CRNs receive auditory descending inputs, we developed tract-tracing studies combined with immunohistochemistry, electron microscopy, morphometry, and confocal microscopy. Either FluoroGold or biotinylated dextran amine (BDA) injections in CRNs showed retrogradely labeled neurons in the ventral nucleus of the trapezoid body (VNTB). We verified the projection to CRNs by injecting BDA into the VNTB. Our results showed that neurons from VNTB project bilaterally and directly to CRNs, giving off numerous endings onto cell bodies and preferentially dendrites of CRNs. Electron microscopy analysis of labeled VNTB terminals demonstrated that they made multiple symmetric synapses and contained small round vesicles. Colocalization of the vesicular acetylcholine transporter and fluorescein dextran after injection in the VNTB indicated that these terminals use acetylcholine as neurotransmitter. We also revealed that the inferior colliculus, an important nucleus mediating the auditory prepulse inhibition, projects to VNTB neurons that innervate CRNs. Our data show a novel and short descending auditory pathway from the VNTB to the first nucleus of the primary acoustic startle circuit that might play an important role in the auditory prepulse inhibition of the startle reflex elicited by sounds.

The genetic audiogenic seizure hamster from Salamanca: The GASH:Sal

“…The animals were euthanized with pentobarbital (60 mg/kg) and perfused transcardially with 0.9% saline wash solution followed by 4% paraformaldehyde fixative solution. Following perfusion, 40-μm coronal serial sections were processed for immunohistochemistry using similar procedures to those used in our previous studies of rats [43] and hamsters [44]. Briefly, the sections were washed and incubated in a rabbit anti-FOS sc-52 primary antibody solution (1:2500; Santa Cruz Biotechnology, Santa Cruz, CA, USA) diluted in TBS (Tris-buffered saline) for 24 h at 4°C.…”

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

“…In another set of experiments, 6 rats received unilaterally injections of BDA into the left DCN and VCN. All surgical and stereotaxic procedures for injecting the BDA were identical to that used in our previous studies (Gómez-Nieto et al, 2008a, 2013; Horta-Júnior et al, 2008). BDA (10% in distilled water) were injected iontophoretically via a glass micropipette (25μm tip diameter), with 3μA positive current pulses (7 s on/7 s off) for a period of 15 min.…”

“…Tissue preparation for light microscopy included: perfusion of the animals, brain dissection and subsequent cryoprotection with sucrose, freezing and serially slicing at 40μm thickness along parasagittal and coronal planes, visualization of HRP and BDA neurotracers, and calbindin protein-D28K (CaBP) immunohistochemistry. All these techniques were applied in a manner identical to that used in our previous reports (Osen et al, 1991; López et al, 1993, 1999; Gómez-Nieto et al, 2008a, 2013). In cases with HRP injections, we followed a standard immunostaining protocol to visualize CaBP as described by Gómez-Nieto et al (2008a).…”

“…All these techniques were applied in a manner identical to that used in our previous reports (Osen et al, 1991; López et al, 1993, 1999; Gómez-Nieto et al, 2008a, 2013). In cases with HRP injections, we followed a standard immunostaining protocol to visualize CaBP as described by Gómez-Nieto et al (2008a). CaBP immunoreactivity has been extensively utilized for detecting the cell body and dendrites of the CRNs (López et al, 1993; Gómez-Nieto et al, 2008a, 2013).…”

“…In cases with HRP injections, we followed a standard immunostaining protocol to visualize CaBP as described by Gómez-Nieto et al (2008a). CaBP immunoreactivity has been extensively utilized for detecting the cell body and dendrites of the CRNs (López et al, 1993; Gómez-Nieto et al, 2008a, 2013). Thus, nickel-intensified peroxidase reaction was developed for HRP visualization in order to distinguish from CaBP immunohistochemistry without heavy metal intensification of diaminobenzidine (Gómez-Nieto et al, 2008a).…”