Donald A. Godfrey

Neurotransmitter Microchemistry of the Cochlear Nucleus and Superior Olivary Complex

The complex function of the nervous system is made possible by its underlying chemistry. One aspect of this chemistry involves the primary mechanism by which neurons communicate with each other, chemical neurotransmission. It is likely that the neurons of the auditory system are organized chemically as well as structurally and functionally. We may therefore look for populations of neurons employing particular neurotransmitters. These neuronal populations may serve specific functions in the auditory system, just as specific functions are served by cholinergic spinal motoneurons or dopaminergic nigrostriatal neurons.

Choline acetyltransferase and acetylcholinesterase in centrifugal labyrinthine bundles of rats

Activities of choline acetyltransferase and acetylcholinesterase were measured for the acetylcholinesterase-positive fiber bundles containing axons projecting from the brainstem to the labyrinth of the rat. These activities were compared to those of a well-established cholinergic tract: the facial motor root. The choline acetyltransferase activities were roughly similar between the tracts, consistent with a conclusion that the centrifugal labyrinthine fibers are all cholinergic. The acetylcholinesterase activities were much higher in the centrifugal labyrinthine bundle than in the facial motor root, probably relating to the smaller diameters of the labyrinthine fibers. Transection of the centrifugal labyrinthine bundle led to virtually total loss of its choline acetyltransferase activity lateral to the cut, consistent with a centrifugal direction of all the fibers, but loss of only half its acetylcholinesterase activity, even after 34 days. These results agree with those for well-established cholinergic pathways, including the facial motor root in the present study, and with previous suggestions that a component of the acetylcholinesterase in cholinergic tracts might be synthesized by cells other than the neurons in the tract.

Effects of trapezoid body and superior olive lesions on choline acetyltransferase activity in the rat cochlear nucleus.

Using a microdissection and quantitative microassay approach, choline acetyltransferase activities were mapped in the cochlear nuclei of rats having either transection of the trapezoid body or destruction of the superior olivary complex on one side in the brain stem. Lateral trapezoid body transection resulted in dramatic loss of choline acetyltransferase activity in all parts of the ipsilateral cochlear nucleus, while more medial transection had little effect. Destruction of the superior olivary complex resulted in dramatic loss of choline acetyltransferase activity in the ipsilateral cochlear nucleus, and detectable loss also contralaterally. The results suggest that most of the centrifugal cholinergic projections to the rat cochlear nucleus derive from or traverse the vicinity of the superior olivary complex bilaterally and enter the cochlear nucleus ventrally from the region of the trapezoid body.

Laminar Distributions of Choline Acetyltransferase and Acetylcholinesterase Activities in the Inner Plexiform Layer of Rat Retina

Abstract: Choline acetyltransferase and acetylcholinesterase activities were measured in samples taken at 7‐μm increments through the inner plexiform layer of rat retina. These enzyme activities were not uniformly distributed through the depth of the inner plexiform layer. Peaks of choline acetyltransferase activity occurred at about onethird and peaks of acetylcholinesterase activity at about one‐fifth of the depth into the inner plexiform layer from either side. The positions of the two peaks of choline acetyltransferase activity most likely correspond to the locations of processes from cholinergic amacrine somata in the inner nuclear layer, which spread in sublamina a, and processes from cholinergic amacrine somata “displaced’ in the ganglion cell layer which spread in sublamina b of the inner plexiform layer. The peaks of acetylcholinesterase activity may in addition correspond to the processes of cholinoceptive amacrine and ganglion cells. The magnitudes of choline acetyltransferase and acetylcholinesterase activities are as high as found anywhere in rat brain, emphasizing the important role of cholinergic mechanisms in visual processing through the rat inner plexiform layer.

Effect of olivocochlear bundle transection on choline acetyltransferase activity in the rat cochlear nucleus

Using a microdissection and quantitative microassay approach, choline acetyltransferase activities were mapped in the cochlear nuclei of rats having complete transections of the olivocochlear bundle on one side in the brain stem. In rats in which the trapezoid body was not significantly damaged by the lesion, consistent reductions of choline acetyltransferase activity in subregions of the lesion-side cochlear nucleus, as compared to the control side, averaged about 20%. Nevertheless, a profound lesion-side reduction of choline acetyltransferase activity was found in a branch connection from the olivocochlear bundle to the cochlear nucleus. The results suggest that branches from the olivocochlear bundle are cholinergic, but contribute a relatively minor proportion of the cholinergic synapses in all regions of the rat cochlear nucleus. In the light of previous results with more extensive lesions, it can be proposed that synapses in all regions of the rat cochlear nucleus. In the light of previous results with more extensive lesions, it can be proposed that most cholinergic input into the rat cochlear nucleus enters by a ventral route along the trapezoid body. It is noted that this represents a quantitatively somewhat different situation from that in the cat.

Effects of large brain stem lesions on the cholinergic system in the rat cochlear nucleus

Large lesions were made medial to one cochlear nucleus in rats, in order to cut virtually centrifugal pathways to it. To estimate the contribution of these centrifugal pathways to cholinergic synapses in the cochlear nucleus, choline acetyltransferase and acetylcholinesterase activities were mapped, by quantitative histochemical procedures, in lesion and control side cochlear nuclei. Choline acetyltransferase activities were reduced by 85-90% in most regions of the lesion side cochlear nucleus and by 65-75% in granular regions. Acetylcholinesterase activities were reduced by 50% or less in the same regions. The choline acetyltransferase results are consistent with a conclusion that by far most cholinergic synapses in the rat cochlear nucleus derive from centrifugal pathways. Additionally, the effects of the lesions on enzyme activities in the lateral superior olivary nucleus and ventral nucleus of the trapezoid body, and in the facial, motor trigeminal, and spinal trigeminal nuclei were examined. In the lesion side facial nucleus, 60% and 40% decreases in choline acetyltransferase and acetylcholinesterase activities, respectively, were apparently consequences of facial root transection. Lesion-control enzyme activity differences in the other nuclei were much smaller.

Separate enzymatic microassays for aspartate aminotransferase isoenzymes

The properties of the cytosolic and mitochondrial isoenzymes of aspartate aminotransferase were studied using a commercial preparation of the cytosolic isoenzyme, a mitochondrial preparation, and whole brain homogenate. Based on these properties, microassays were developed and shown to be highly specific and quantitatively accurate for measuring the activity of either the cytosolic or mitochondrial isoenzyme in microgram quantities of tissue. The assays have been successfully applied to homogenates of a wide variety of tissues. They can be used to measure the activities of aspartate aminotransferase isoenzymes in sub-microgram samples of freeze-dried tissue.

Contribution of centrifugal innervation to choline acetyltransferase activity in the cat cochlear nucleus

Using a quantitative microchemical mapping approach combined with surgical cuts of fiber tracts, the contributions of centrifugal pathways to choline acetyltransferase activity were mapped three-dimensionally in the cat cochlear nucleus. Large reductions of choline acetyltransferase activity, averaging 70%, were measured in almost all parts of the lesion-side nucleus following transection of virtually all its centrifugal connections. More superficial cuts, penetrating just through the olivocochlear bundle, also led to significant reductions of enzyme activity, especially most rostrally in the anteroventral cochlear nucleus and superficial granular region, where the reductions were similar to those following the complete cuts. Lesions encroaching upon the superior olivary complex gave bilateral effects. Transverse cuts between rostral and caudal parts of the cochlear nucleus gave some small effects. The results suggest that, as in rats, most choline acetyltransferase activity in the cat cochlear nucleus is associated with its centrifugal innervation. However, unlike the situation in rats, the enzyme activity in cats is related more to olivocochlear branches than to ventral fibers in the trapezoid body region. Also, the choline acetyltransferase activity related to olivocochlear collateral innervation is much less uniformly distributed within the cochlear nucleus in cats than in rats.

Aspartate Aminotransferase Activity in Fiber Tracts of the Rat Brain

Abstract: Activity of aspartate aminotransferase, an enzyme which catalyzes the interconversion of the excitatory transmitter candidates, glutamate and aspartate, has been measured in fiber tracts of rat, with an emphasis on sensory and motor systems of the brain. Most tracts had significantly higher activities than the cholinergic facial nerve root, consistent with the possibility that a component of aspartate aminotransferase activity might serve as a marker for neurons using glutamate and/or aspartate as neurotransmitter. Highest activity was in the auditory nerve root. On the other hand, a close correlation was found between aspartate aminotransferase and malate de‐hydrogenase activities in the fiber tracts, raising the question whether aspartate aminotransferase activity may be more closely related to energy metabolism than to transmitter metabolism.

Quantitative distributions of aspartate aminotransferase and glutaminase activities in the rat cochlea

The intra-cochlear distributions of aspartate aminotransferase and glutaminase, prominent enzymes of aspartate and glutamate metabolism, have been studied by quantitative microchemical techniques. Also measured was choline acetyltransferase, the enzyme synthesizing acetylcholine, and a marker for the olivocochlear bundle. Aspartate aminotransferase activity was highest in the stria vascularis, about half this high in the organ of Corti synaptic (hair cell) zones, somewhat lower in the organ of Corti non-synaptic (Hensens cell) zones, lower yet in Reissners and lowest in the tectorial membrane. Glutaminase, on the other hand, had its highest activity in synaptic zones, about a third of that activity in the organ of Corti non-synaptic zones, and a barely detectable activity in Reissners and tectorial membranes, and stria vascularis. Seven days after transection of the olivocochlear bundle, no significant difference was found between lesion- and control-side aspartate aminotransferase or glutaminase activities, even though no choline acetyltransferase activity remained in the lesion-side of the organ of Corti. Both the distribution of aspartate aminotransferase activity and the lesion results would seem to implicate it in energy more so than neurotransmitter metabolism. The distribution of glutaminase activity could be consistent with a role in neurotransmission; however, the lesion data were unable to demonstrate a specific association with the olivocochlear bundle.