James C. Hazlett

GABA-immunoreactivity in ganglion cells of the rat retina☆

Ganglion cells in the rat retina were labeled with the fluorescent dye, Diamidino-yellow, by retrograde transport from the superior colliculus and subsequently reacted for GABA-like immunoreactivity with a rhodamine-conjugated antiserum. Examination of sectioned retinas by fluorescence microscopy showed double labeling in approximately 6% of the ganglion cells. The presence of GABA in these neurons suggests that they may be involved in providing direct inhibitory input to the rat tectum.

Immunocytochemical evidence for the involvement of glycine in sensory centers of the rat brain.

Glycine-like immunoreactivity was localized to a number of sites in the rat brain which are involved in processing sensory information. In the auditory and vestibular systems, glycine immunoreactivity was seen in dorsal and ventral cochlear nuclei, superior olive, trapezoid body, medial and lateral vestibular nuclei, and inferior colliculus. Staining in the visual system was seen in retina, dorsal lateral geniculate nucleus, and superior colliculus. The olfactory system exhibited staining in the olfactory bulb and accessory olfactory formation. Somatosensory centers with glycine immunoreactivity included the dorsal column nuclei, spinal trigeminal nucleus, principal sensory nucleus of V, reticular formation, and periaqueductal gray. Glycine-immunoreactive neurons were also seen in cerebellar cortex, deep cerebellar nuclei, hippocampus, cerebral cortex, and striatum. The distribution of staining indicates that glycine plays a major role in sensory centers with actions at both strychnine-sensitive and strychnine-insensitive receptors.

KINETICS OF ENTRY OF PROTEINS INTO THE MYELIN MEMBRANE

Brain slices were prepared from 17‐day old rats, and incubated with [3H]glycine or [3H]‐leucine to label proteins. Myelin was isolated from the slices, and the proteins were separated by discontinuous gel electrophoresis in buffers containing sodium dodecyl sulfate. Radioactive basic and Wolfgram proteins appeared in myelin at similar initial rates, and their entry was nearly linear between 15 and 120 min with no detectable lag. Radioactive proteolipid protein appeared in myelin at one‐fourth the rate of the basic and Wolfgram proteins between 0 and 30 min, then entered at a rate comparable to the other proteins between 45 and 120 min.

Observations on the early development of ascending spinal pathways

SummaryThe development of ascending spinal pathways has been studied in the North American opossum using degeneration methods and the retrograde transport of horseradish peroxidase. Axons from caudal thoracic and/or lumbosacral levels of the spinal cord reach the lateral reticular nucleus, the inferior olivary complex, the reticular formation of the medulla and pons as well as the cerebellum very early in development. Innervation of the nucleus gracilis occurs somewhat later. Spinal axons grow into most of the caudal brain stem areas they occupy in the adult animal, including the nucleus gracilis, before there is convincing evidence that they reach the thalamus. Although spinal axons enter the cerebellum early in development their adult distribution with its characteristic discontinuities appears relatively late.

Origin and topography of thalamocaudate projections in the opossum

The origins of the thalamocadaute projections in the North American opossum include the intralaminar nuclei (parafascicular complex, the paracentral, centralis and central lateral nuclei) and the intermedial dorsal, medial dorsal and interanterior dorsal nuclei. Of these, portions of the intralaminar nuclei exhibit rudimentary elements of a topographically organized projection. However, it is obvious that the patterns observed here are much less specific than those reported in other mammalian species. We suggest that the question of topographic specificity may relate to the overlapping of sensory and motor representations in the neocortex.

The development of selected rubral connections in the North American opossum

We have employed axonal transport and degeneration techniques to study the development of selected rubral connections in the North American opossum. Opossums were chosen for study because they are born in an immature state, 12 days after conception, and have a lengthy postnatal development. The results of our studies suggest that: (1) the red nucleus innervates the spinal cord early in development, but not as early as some areas of the brainstem; (2) rubrospinal development occurs postnatally in the opossum; (3) rubrospinal axons do not grow synchronously into the spinal cord, but are added over time; (4) rubrospinal development follows rough rostral to caudal and lateral to medial gradients; (5) the red nucleus is innervated by the cerebellum well before it receives projections from the cerebral cortex; and (6) cortical axons do not grow into the red nucleus until after rubrospinal axons have reached most of their adult targets.

Development of the basilar pons in the North American opossum: dendrogenesis and maturation of afferent and efferent connections

SummaryThe present study provides data on temporal factors that may play a role in the development of precerebellar-cerebellar circuits in the North American opossum. In this study the basilar pons and cerebellum are analyzed from birth, 12–13 days after conception, to approximately postnatal day (PD) 80 at which time the brainstem and cerebellum have a mature histological appearance. In Nissl preparations, the basilar pons was first seen at PD 7 as a small cluster of tightly packed cells. Analysis of Golgi impregnations revealed that dendritic growth occurred between PD 25–80. During this period, dendrites gradually increased in length and in the complexity of their branching pattern. Horseradish peroxidase (HRP) was placed into the cerebellar and cerebral cortices in order to examine the development of efferent and afferent projections of the basilar pons, respectively. Evidence for the growth of pontine axons into the cerebellum was first detected on PD 17. Neurons located dorsally within the basilar pons appear to be the first neurons retrogradely labeled with horseradish peroxidase. By PD 27 retrogradely labeled neurons are found throughout the basilar pons. Afferent fibers from the cerebral cortex are not seen within the neuropil of the nucleus until after PD 25 and by PD 29, they have greatly expanded their terminal fields. Degeneration techniques reveal that afferent fibers from the cerebellum arrive by PD 19 and increase in number until PD 30 when their adult distribution is achieved. These data suggest that the time of afferent arrival from the cerebral cortex and deep cerebellar nuclei is closely correlated in time with the initiation of dendritic maturation and the outgrowth of pontocerebellar axons. Afferent axons from the cerebral cortex and deep cerebellar nuclei reach the basilar pons and afferents from the basilar pons grow into the cerebellum when the dendrites of the respective target neurons are very immature. Thus, the time of axon arrival in these circuits may be an important factor in determining their synaptic location on individual neurons. The data derived from the present study is compared to those obtained in previous studies on the inferior olive. The results of this comparison provide evidence for a similar sequence of events, but a differential timetable for the development of specific connections within precerebellar-cerebellar circuits.

The early development of major projections to the dorsal striatum in the North American opossum

We have employed immunocytochemical and axonal transport techniques to study the development of major projections to the dorsal striatum of the North American opossum. The opossum is born in a very immature state, 12-13 days after conception, and climbs into an external pouch where it remains attached to a nipple for several months. Its immaturity at birth and its protracted postnatal development make the opossum a good model for developmental studies. Although tyrosine hydroxylase-like immunoreactive (TH-LI), presumably dopaminergic, neurons were present in the ventral mesencephalon at birth (the presumptive substantia nigra and ventral tegmental area), there was no evidence for TH-LI axons in the striatal anlage. By postnatal day (PD)6, a few immunostained axons were found within the putamen. The subsequent growth of TH-LI axons into the striatum followed general caudal to rostral and ventrolateral to dorsomedial gradients and, at any age, they were most numerous in the areas exhibiting the greatest cytodifferentiation. By estimated (E)PD45, TH-LI axons were present in most, if not all, areas of the striatum. Serotoninergic (5-HT)-LI axons were found lateral to the presumptive striatum at birth but not within it. By PD7, however, a few 5-HT-LI axons could be identified in the putamen. The growth of 5-HT-LI axons into the striatum generally followed the same gradients described for TH-LI axons although at all ages their density was much less. Using the orthograde transport of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP), evidence was obtained for the existence of thalamostriatal projections by PD5 and for corticostriatal projections by PD10. Crossed corticostriatal projections were present by EPD23. Our results suggest that the development of major projections to the striatum occurs postnatally in the opossum, rather than prenatally as in placental animals. The timetable for striatal innervation is discussed in light of the developmental sequences established for other motor circuits.

Cerebrovascular responses to pentylenetetrazol: time and dose dependent effects

The effects of subconvulsant and convulsant doses of pentylenetetrazol (PTZ) on cerebral blood flow (rCBF), permeability-capillary surface area products (rPS), and brain vascular spaces (BVS) were examined in 15 brain regions at 1 h, 24 h and 1 week after injection in male Sprague-Dawley rats. Brain histology was examined 3 days after injection. A dose of PTZ (50 mg/kg, i.p.), sufficient to trigger a single convulsive seizure, produced small regional changes in rCBF at 1 h, but not at 24 h or 1 week after injection. No significant changes in rPS or BVS were found at any time, and only mild histologic changes were observed. In contrast, a dose of PTZ (25 mg/kg) which failed to cause either convulsions or significant electrocorticographic changes, markedly increased rCBF and rPS. Some of these regional effects were still observed 1 week later. Similarly, more severe and extensive cellular changes followed treatment with the subconvulsive dose. These findings indicate that PTZ treatment can have prolonged effects on cerebrovascular functions and neuronal integrity even in the absence of convulsive activity.

Innervation of the pituitary gland by supraependymal neurons

Retrograde transport of horseradish peroxidase (HRP) was used to identify supraependymal neurons projecting to the pituitary gland in the hamster. Supraependymal neurons overlying the median eminence were labeled by HRP injections into the neural and intermediate lobes of the pituitary gland; no neurons were labeled following HRP injections confined to the anterior lobe. Supraependymal neurons innervating the pituitary gland may provide a means whereby cerebrospinal fluid-borne substances modulate neuro-intermediate lobe function.