Effects of surgical lesions on choline acetyltransferase activity in the cat cochlea.

Although it is well established that the choline acetyltransferase (ChAT, the enzyme for acetylcholine synthesis) in the mammalian cochlea is associated with its olivocochlear innervation, the distribution of this innervation in the cochlea varies somewhat among mammalian species. The quantitative distribution of ChAT activity in the cochlea has been reported for guinea pigs and rats. The present study reports the distribution of ChAT activity within the organ of Corti among the three turns of the cat cochlea and the effects of removing olivocochlear innervation either by a lateral cut aimed to totally transect the left olivocochlear bundle or a more medial cut additionally damaging the superior olivary complex on the same side. Similarly to results for guinea pig and rat, the distribution of ChAT activity in the cat outer hair cell region showed a decrease from base to apex, but, unlike in the guinea pig and rat, the cat inner hair cell region did not. As in the rat, little ChAT activity was measured in the outer supporting cell region. As previously reported for whole cat cochlea and for rat cochlear regions, transection of the olivocochlear bundle resulted in almost total loss of ChAT activity in the hair cell regions of the cat cochlea. Lesions of the superior olivary complex resulted in loss of ChAT activity in the inner hair cell region of all cochlear turns only on the lesion side but bilateral losses in the outer hair cell region of all turns. The results are consistent with previous evidence that virtually all cholinergic synapses in the mammalian cochlea are associated with its olivocochlear innervation, that the olivocochlear innervation to the inner hair cell region is predominantly ipsilateral, and that the olivocochlear innervation to the outer hair cells is bilateral.

Inhibition of MMP-2 but not MMP-9 influences inner ear spiral ganglion neurons in vitro.

Matrix metalloproteinases (MMPs) play an important role in modeling of the extracellular matrix. There is increasing evidence that these proteases are important in neurite elongation and axonal guidance during development in the central nervous system and retina. Moreover, they are also expressed after acute injury and can be the key mediators of pathogenesis. However, the role of MMPs in the inner ear is largely unknown. Our group recently demonstrated that general inhibition of MMPs resulted in auditory hair cell loss in vitro. In the present study, we investigated the role of MMPs in inner ear spiral ganglion neuron (SGN) survival, neuritogenesis and neurite extension by blocking MMPs known to be involved in axonal guidance, neurite elongation, and apoptosis in other neuronal systems. Spiral ganglion (SG) explants from 5-day-old Wistar rats were treated with different concentrations of the general MMP inhibitor GM6001, a specific MMP-2 inhibitor, and a specific MMP-9 inhibitor, in vitro. The general inhibitor of MMPs and the specific inhibition of MMP-2 significantly reduced both the number of neurites that extended from SG explants, as well as the length of individual neurites. However, neither the general inhibitor of MMPs nor the specific inhibition of MMP-2 influenced SGN survival. Inhibition of MMP-9 had no influence on SGNs. The data suggest that MMPs, and more specifically MMP-2, influence the growth of developing afferent neurites in the mammalian inner ear in vivo.

Distribution of the Na,K-ATPase alpha subunit in the rat spiral ganglion and organ of corti.

Processing of sound in the cochlea involves both afferent and efferent innervation. The Na,K-ATPase (NKA) is essential for cells that maintain hyperpolarized membrane potentials and sodium and potassium concentration gradients. Heterogeneity of NKA subunit expression is one mechanism that tailors physiology to particular cellular demands. Therefore, to provide insight into molecular differences that distinguish the various innervation pathways in the cochlea, we performed a variety of double labeling experiments with antibodies against three of the alpha isoforms of the NKA (NKA alpha 1-3) and markers identifying particular subsets of neurons or supporting cells in whole mount preparations of the organ of Corti and spiral ganglion. We found that the NKA alpha 3 is abundantly expressed within the membranes of the spiral ganglion somata, the type I afferent terminals contacting the inner hair cells, and the medial efferent terminals contacting the outer hair cells. We also found expression of the NKA alpha 1 in the supporting cells that neighbor the inner hair cells and express the glutamate transporter GLAST. These findings suggest that both the NKA alpha 1 and NKA alpha 3 are poised to play an essential role in the regulation of the type I afferent synapses, the medial efferent synapses, and also glutamate transport from the afferent-inner hair cell synapse.

Dense innervation of Deiters' and Hensen's cells persists after chronic deefferentation of guinea pig cochleas.

Innervation of Deiters' and Hensen's cells has been described in the organ of Corti of several mammalian species and has been suggested to arise from the olivocochlear (OC) efferent system (Wright and Preston [1976] Acta Otolaryngol. 82:41-47). In the present study, antineurofilament immunostaining was used to reveal these outer supporting cell fibers (OSCFs) in the normal guinea pig. In control ears, OSCFs were absent in the basal half of the cochlea but increased in number steadily toward the apex, peaking at values of over 1,200 fibers/mm. These values indicate a far more profuse innervation of supporting cells than has been described previously, suggesting that most OSCFs were not stained in previous immunohistochemical studies. Chronic cochlear deefferentation was used to test whether OSCFs are part of the OC system. The OC bundle was transected unilaterally, and the animals were allowed to survive for 4-8 weeks. Completeness of deefferentation was assessed by using acetylcholinesterase staining of the brainstem and measurement of the density of OC fascicles in the cochlea. By using these metrics, unilateral deefferentation was nearly complete in three animals. In successfully deefferented cases, the OSCF innervation density was not statistically different from control values. We conclude that the vast majority of OSCFs are not of OC origin. We speculate that they may be branches of type II afferent fibers to outer hair cells and that a smaller population of OSCFs with different morphology and immunoreactivity may arise from the OC system.