Neurotrophin expression of laryngeal muscles in response to recurrent laryngeal nerve transection.

OBJECTIVE/HYPOTHESIS: The recurrent laryngeal nerve (RLN) commonly regenerates after injury; however, functional motion is rarely recovered. Animal experiments have documented aberrant reinnervation after nerve transection, with motor axons reaching inappropriate muscles. More recently, experimental results suggest that lack of vocal fold motion after RLN injury is due to preferential reinnervation of adductor muscles, with inadequate reinnervation of the posterior cricoarytenoid muscle (PCA), the only abductor muscle of the larynx. Information on factors that could influence the receptiveness of these muscles to reinnervation could be useful in developing new therapeutic strategies. It is hypothesized that the thyroarytenoid muscle (TA) and the PCA differ in expression of neurotrophins in response to denervation. STUDY DESIGN: Laboratory experiment. METHODS: Rats were sacrificed at 3 days, 6 weeks, or 4 months after unilateral RLN injury measure expression of brain-derived nerve growth factor (BDNF), nerve growth factor (NGF), and neurotrophin 4 (NT-4) in the TA and PCA muscles, using immunohistochemistry. We also assessed nerve regeneration. RESULTS: NGF was significantly diminished in the denervated TA muscle at 3 days after injury and increased at 6 weeks. BDNF expression was unchanged in the TA, but was diminished in both PCA muscles at 3 days and 6 weeks, returning to near-normal levels at 4 months after injury. Robust nerve regeneration of distal RLN was present at 4 months. CONCLUSIONS: Results suggest that the TA and PCA muscles respond differently to denervation.

A classification of NOergic neurons in the inferior colliculus of rat according to co-existence with classical amino acid transmitters.

Since the localization of nitric oxide synthase (NOS) can be identified by enzyme histochemistry for NADPH-diaphorse (NADPH-d), this method has been used widely for mapping NOS-containing (presumably NOergic) neurons in the central nervous system. So far several studies suggest that NADPH-d is present in distinct neuronal populations in the inferior colliculus (IC), a major processing center for both the ascending and descending auditory pathway, and NO may play an important role in audition. On one hand, there is evidence from several lines of research that the IC makes extensive use of the neuroactive amino acids, in particular the inhibitory transmitter g-aminobutyric acid (GABA) and the excitatory amino acid glutamate (GLU). However, lacking is a description of the distribution of NOergic neurons to which traditional neurotransmitters may be linked. The present research utilized NADPH-d enzyme histochemistry in combination with immunocytochemistry to determine if NO may colocalize with either or both GABA and glutamate in distinct subpopulations of IC neurons. The NADPH-d positive neurons were predominantly found in two main subdivisions of the IC: the external cortex (ECIC) and the dorsal cortex (DCIC). The large numbers of these NADPH-d positive neurons appeared immunostained for GLU while only a small number, seemed to belong to the small cells (somatic area < 100 microm2) similarity to stellate cells group was positive for GABA throughout the cortex of the IC. Owing to no coexistence between GABA and GLU in the same NADPH-d positive neuron in the pairs of adjacent sections of the IC by the mirror-image technique, the present results consequently support that NOergic neurons could be subdivided into at least three distinct populations with a large proportion of about 77% being GLUergic, much lower frequency of about 11% being GABAergic and the remaining 12% expressing non-GABA and non-GLU. In summary, the existence of two functionally distinct populations of NO/GABAergic and NO/GLUergic neurons in the NOergic neurons of IC suggest that at least two differential pattern of GLU-mediated excitatory NO transmission and GABA-mediated inhibitory NO transmission are involved in the networking of auditory communication in the cortex of IC.

Age-related synaptic changes in the anteroventral cochlear nucleus of Fischer-344 rats.

Previous studies have demonstrated age-related decreases in the transmitters glycine and glutamate in the cochlear nucleus (CN) of the Fischer-344 (F344) rat, along with declining levels of binding for glycine receptors. The purpose of this study was to evaluate structural correlates to the transmitter and receptor losses that accompany aging in the anteroventral CN (AVCN). Thin sections were obtained from the middle-frequency area of the right AVCNs from five 3-month-, four 19-month-, and five 28-month-old F344 rats. Montages were constructed from electron micrographs taken of several sites in each AVCN section. The presynaptic terminals were classified by vesicle type and postsynaptic target, and their perimeters and synaptic lengths were traced using morphometry software. The calibers of all dendritic profiles were also measured, and cell counts were performed on semi-thin sections. The data were compared among the three age groups using analysis of variance followed by Tukey's Honestly Significant Difference for pairwise comparisons. There were significant age-related decreases in the size of terminals contacting small-caliber (<2 microm) dendrites. Dendrites of this size comprised the largest percentage of dendrites in the AVCN. On these targets, round and pleomorphic-vesicle terminals were reduced in volume by nearly 44% and 24%, respectively, in 28-month olds when compared to the 3-month olds. On the other hand, the densities and numbers of synaptic terminals and dendritic profiles did not differ among age groups, and no neuronal losses were evident in the older animals. Also, there were no detectable changes in synaptic area among groups. The decrease in terminal size may be related to age-associated reductions in neurotransmitter levels previously described in the F344 CN. The observations presented here contrast with those previously described in the inferior colliculus (IC), in which there were significant age-related losses of synaptic terminals and dendrites, but no change in the size of synaptic terminals. The lack of synaptic and dendritic losses suggests that the structural connectivity of the rat AVCN remains relatively intact during aging, which is interesting in light of the synaptic and dendritic changes evident in the IC, a major target of its projections.

Hsp70 in the inferior colliculus of Fischer-344 rats: effects of age and acoustic stress.

Heat shock proteins 72 and 73 (hsp72 and hsp73) were studied in the inferior colliculus (IC) of Fischer-344 rats to determine if their levels are altered during normal aging and following exposure to intense acoustic noise. Three age groups of rats (3, 18, and 25 months) were exposed to ambient sound (control) or broad-band noise at 108 dB sound pressure level (0.0004 dyn/cm2) for 30 min. Western blotting procedures were used to measure hsp72 and hsp73 in ICs and cerebella (positive control). Immunohistochemistry was performed using 3-month olds to study the localization patterns of hsp72 and hsp73 in both structures. The IC and cerebellum exhibited immunolabeling over neuronal somata and proximal dendrites. Ambient levels of hsp72 in supernatants from aged rats were reduced 56.5%+/-7.8% in the IC relative to 3-month olds. This decrease may render the IC more susceptible to stress-related damage. An increase in constitutive hsp73 (350.7%+/-70.4%) was observed in IC pellet fractions from animals exposed to the 108-dB noise when compared to the ambient-noise controls, suggestive of a lipoprotective role for hsp73. This elevation was consistent across age groups. No noise-induced changes in hsp72 were detectable in the IC, indicating that loud sounds may not be an appropriate stimulus for hsp72 induction in this structure.