Applications of autoradiography in neurobiological research.
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Amino Acids↗
Biomedical subjects
C Pilgrim
Publications and source records attributed to C Pilgrim.
Origin and terminals of the pretectothalamoclaustral connections in the cat.
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Afferent Pathways↗
Distribution of thyrotropin-releasing hormone (TRH) immunoreactivity in the thoracic spinal cord of guinea pig.
The localization of thyrotropin-releasing hormone immunoreactivity (TRH-IR) has been determined in the thoracic spinal cord of male and female guinea pigs. The immunoreactive product is localized in nerve fibre varicosites and terminals throughout the spinal gray matter and in some regions of the white matter. There are not TRH-IR neurons in the spinal cord. The highest density of IR structures is observed in the intermediate zone, in the central gray and in the ventral horns, around the motoneurons. Less TRH-IR structures are observed in the superficial layers and substantia gelatinosa of the dorsal horns. Between the ependymal cells of the central canal are observed single TRH-IR fibres and terminals too. Most of the TRH-IR fibres and terminals in the intermediate zone and in the central gray are constituents of the vegetative network (Galabov and Davidoff, 1976 and Davidoff et al. 1985). As to the origin of the spinal cord TRH-IR fibres and terminals two main possibilities exist: a) From primary afferent neurons situated in the dorsal root ganglia, which is quite uncertain; b) From supraspinal neurons which send their axons descending in the white matter and in the fasciculi longitudinales laterales and mediales (Johansson et al. 1981, 1983). The wide diversity of neuroactive substances in the thoracic spinal cord vegetative network and the origin of its fibres and terminals suggests that this network may play an important role in the integration of the central and peripheral vegetative nervous system, as well as probably in the integration of the somatic and the vegetative nervous system.
Animals↗
Projections from the pretectothalamic border area to the somatosensory part of the claustrum: an autoradiographic study.
Injections of tritiated amino acid into the pretectothalamic border area of cats resulted in the appearance of numerous silver grains in the ipsilateral claustrum. The accumulation of grains was especially abundant in the dorsomedial area of the central part of the claustrum. These results indicate the presence of substantial projection from the pretectothalamic border area to the claustrum. It may be assumed that these projections belong to the somatosensory system, since they connect mainly di(mes)encephalic nuclei, receiving largely the somatosensory input, with the somatosensory part of the claustrum.
Amino Acids↗
Retrograde transport of fluorescent tracers reveals extensive ipsi- and contralateral claustrocortical connections in the rat.
The projections from the claustrum to the cerebral cortex in the rat were examined by means of retrogradely transported fluorescent tracers Fast Blue (FB) and Diamidino Yellow dihydrochloride (DY), injected in the prefrontal, motor, somatosensory, auditory, and visual fields. In all cases, substantial numbers of retrogradely labeled neurons were observed in the ipsilateral and moderate to scant numbers in the contralateral claustrum insulare. Symmetrical bilateral injections of FB and DY as well as simultaneous injections of the tracers in the motor and visual cortex of the same hemisphere revealed no double-labeled neurons in the claustrum. The following conclusions may be drawn: The claustral projections to the motor, somatosensory, and visual cortex are prominent. The projection to the prefrontal cortex is less substantial and that to the auditory cortex is relatively modest. The claustrocortical connections lack the clear-cut topographic pattern of the thalamic nuclei but are, to some degree, preferentially arranged, albeit with considerable overlapping of the subpopulations of corticopetal neurons, a coarse anteroposterior topographic distribution appears to exist also in rodents. Neurons contributing to the claustrocortical connection project either ipsilaterally or contralaterally but not bilaterally. Projections to different cortical fields of one hemisphere also originate from separate claustral neurons.
Animals↗
Diencephalic and mesencephalic afferents of the rat claustrum.
The retrograde fluorescent tracers Fast Blue (FB) and Diamidino Yellow (DY) have been used to study subcortical afferents of the claustrum. DY or FB was injected into the claustrum. The greatest amount of labeled cell bodies were observed in the posterior thalamic nuclear complex. They were especially abundant in its caudal part, lying between the medial geniculate body and the pretectal area. In comparison to the numerous labeled cells near the diencephalic-mesencephalic junction, the number of fluorescing neurons in the brain stem was considerably lower. These neurons were mostly concentrated in the monoaminergic cell groups. The results indicate the presence of a substantial projection from the posterior thalamic and anterior pretectal region to the claustrum.
Afferent Pathways↗
High resolution autoradiographic determination of the topographic distribution of radioactivity in the hippocampal formation after injection of [1-14C]glucose or 2-deoxy[14C]glucose.
Using high resolution autoradiography, the accumulation of radioactivity after intravenous injection of [1-14C]glucose was measured in the corpus callosum, hippocampus, dorsal hippocampal commissure, somatosensory cortex, inferior colliculus and pontine periaqueductal grey. Autoradiograms were prepared by thaw-mounting 4 micron frozen sections on nuclear emulsion-coated slides, and were evaluated quantitatively with a computer-assisted video system for automated counting of silver grains. In all brain regions examined, silver grain densities were greater in rats killed 30 min after injection of [1-14C]glucose compared to rats killed 10 min after injection. After intravenous injection of [1-14C]glucose or 2-deoxy[14C]glucose, the relative uptake and retention of radioactivity in different hippocampal subregions was compared. Striking differences were found in the hippocampus between 2-deoxy[14C]glucose and [1-14C]glucose autoradiograms. After injection of 2-deoxy[14C]glucose, there were large variations in the uptake and retention of radioactivity among different pyramidal cell fields. The CA 3 pyramidal cell field retained considerably more radioactivity than other pyramidal cell fields after injection of 2-deoxy[14C]glucose, while after injection of [1-14C]glucose, the retention of radioactivity was similar in all pyramidal cell fields. After [1-14C]glucose injection, the dentate gyrus contained relatively high levels of radioactivity and more 14C accumulated in the granular layer, compared to the molecular layer. In contrast, after 2-deoxy[14C]glucose injection, there was uniformly less radioactivity throughout the dentate gyrus when compared to rats injected with [1-14C]glucose and there was no preferential accumulation of 2-deoxy[14C]glucose in the granular layer compared to the molecular layer.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Different developmental schedules of dopaminergic and noradrenergic neurons in dissociation culture of fetal rat midbrain and hindbrain.
The development of dopaminergic and noradrenergic neurons in dissociation cultures of mesencephalon and rhombencephalon obtained from 18-day-old rat fetuses was characterized by their capacity to take up and release catecholamines. In both types of cultures, uptake of [3H]dopamine and [3H]noradrenaline was obtained which could be inhibited by reserpine. Autoradiographic studies demonstrated an almost exclusive neuronal localization of the labeled catecholamines. The transmitters could be released by depolarization with K+ in a Ca2+-dependent manner during the entire cultivation period. In contrast, catecholamine uptake by cultures of neocortex was minimal, could not be inhibited by reserpine, and the accumulated radioactivity could not be released upon depolarization. These points provide evidence for an active accumulation of the exogenous transmitters and for the presence of stimulus-secretion coupling in a distinct population of neurons of both brain stem cultures. Striking differences between the two brain stem cultures concerned their sensitivity to desmethylimipramine and benztropine as well as the time course of the development of the uptake capacity. Desmethylimipramine inhibited the uptake of both catecholamines in rhombencephalic, but not in mesencephalic cultures. The reverse was true for benztropine. It is concluded that cultures of rhombencephalon contain predominantly noradrenergic, and those of mesencephalon dopaminergic cells. Comparison of the uptake behaviour suggested that noradrenergic neurons mature considerably later than dopaminergic neurons. The results show that dissociation cultures of mid- and hindbrain, inspite of their heterogeneous composition, can serve as valuable models for the study of development and function of dopaminergic and noradrenergic neurons, respectively.
Adrenergic Fibers↗
Developmental action of a neuronal peptide on a peptidergic neuron.
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Angiotensin II↗
Evidence for an anterograde transport of the fluorescent tracer Fast Blue in the strio-nigro-striatal loop of the rat.
The report is concerned with serendipitous observations which were made in the course of investigations on the connectivity of the claustrum. Stereotaxical injections of Fast Blue were made in the neostriatum. All major sources of striopetal fiber systems were found to contain fluorescent cell bodies. Quite unexpectedly, also fluorescent axons and terminals could be observed in brain regions known to receive a striofugal input including the globus pallidus, entopeduncular and subthalamic nucleus, and the substantia nigra, zona reticulata. It is concluded that Fast Blue may be used for both retrograde and anterograde neuronal tracing.
Amidines↗
Efferent connections of the claustrum to the posterior thalamic and pretectal region in the rat.
Injection of the fluorescent retrograde tracers Fast Blue and Diamidino Yellow in the posterior thalamic and anterior pretectal region of rats resulted in significant retrograde neuronal labeling of the ipsilateral claustrum. The labeled cell bodies were especially abundant in the central third of the claustrum, while the rostral and caudal portions of the nucleus contained only few fluorescing cells. Thus, in addition to the major reciprocal connections of the claustrum with the cerebral cortex, a substantial number of claustral neurons project to the posterior thalamic and anterior pretectal region. Unlike the cortico-claustrocortical loop, this connection appears to be only ipsilateral.
Amidines↗
Quantitative assessment of GABA-uptake sites in the neural lobe by electron-microscopic autoradiography.
Distribution of (3H)GABA in the rat neural lobe was investigated 5 min after intracarotid administration using quantitative electron-microscopic autoradiography. Specificity of (3H)GABA-uptake was tested by pretreatment of control animals with nipecotic acid. It was concluded that, apart from a small fraction in the perivascular spaces, radioactivity was present exclusively in pituicytes. The results confirm and quantify earlier in-vitro observations; they are compared with recent immunocytochemical findings that reveal the presence of glutamate-decarboxylase-containing axons in the neural lobe. It is concluded that there may be GABAergic terminals that lack an uptake mechanism for exogenous transmitter. Nevertheless, (3H)GABA autoradiography is useful in demonstrating other functional components of GABAergic systems, i.e., glial cells.
Animals↗
Hypothalamo-neurohypophysial system in hagfish: localization of neurophysin-, arg-vasopressin- and oxytocin-like immunoreactivity.
Serial paraffin sections of hagfish brain (Eptatretus burgeri) were immunocytochemically examined for porcine neurophysin (NEU), arg-vasopressin (AVP) and oxytocin (OT). A big number of NEU-immunoreactive nerve cells were visible in the frontal part of the ventromedial hypothalamus in close vicinity of the third ventricle. These cells were bipolar in most cases and sometimes additionally stained for AVP but not for OT. NEU-reactive neurons extended axons to the median eminence and the posterior lobe of the pituitary which also contained AVP or OT-like immunoreactivity. Immunostaining outside the hypothalamus could not be observed. It is likely that there exists a mammalian-like oxytocinergic and a vasopressinergic hypothalamo-neurohypophysial system in hagfish, despite the large differences in levels of organization.
Animals↗
The glial reaction in the course of axon regeneration: a stereological study of the rat hypoglossal nucleus.
Both hypoglossal nuclei were examined by electron microscope stereology after unilateral axotomy. The principal aim of this study was a quantitative assessment of the accompanying glial reaction. Volume densities (%) of neuronal and glial perikarya, as well as their processes, were evaluated in terms of volume plus surface densities (mm-1). In addition, specific surfaces (surface to volume ratio) of these neuronal and glial processes were assessed. First, a temporary decrease of dendritic volume density was detected on the ipsilateral side only. Further, the astrocytic reaction displayed differences between stem and lamellar processes. One day after axotomy, a bilateral decrease of volume density, as well as surface density of stem processes, was observed, yet their normal dimensions soon were reestablished. However, a more severe lamellar process reaction was evident. During the first 4 days, a significant increase of volume density and surface density was apparent. In the contralateral hypoglossal nucleus, this glial reaction also occurred but disappeared by day 14, whereas the ipsilateral nucleus continued to display a severe reaction of lamellar processes, only returning to normal status at day 77. In addition, a transient, severe reaction of presumptive microglia was established by employing the volume density and surface density quantitation procedure. Nevertheless, in comparison with the volume and surface contribution of astrocytic processes, the presumed microglial component was minimal. This study indicates a two-step involvement of astrocytes in regenerative repair. Namely, the first phase seems to result in an increase of lamellar processes through reshaping of the stem process.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Claustro-neocortical connections in the rat as demonstrated by retrograde tracing with Lucifer yellow.
The retrograde fluorescent tracer Lucifer Yellow (L.Y.) has been used to study claustro-neocortical connections in the rat with respect to their topography. L.Y. was injected in different cortical fields. Labeled cells were found in the ipsilateral claustrum. The distribution of labeled cells reflects the position of the injection sites along the rostro-caudal axis. The results demonstrate that claustro-neocortical projections are arranged in a topographical fashion not only in the cat, rabbit and monkey, but also in the rat. The connections are not as numerous as in the cat and are only ipsilateral. Our study shows in addition that L.Y. can be used as a connectivity tracer in the nervous system.
Animals↗
Coexistence of glutamate decarboxylase and somatostatin immunoreactivity in cultured hippocampal neurons of the rat.
The immunoperoxidase-antiperoxidase method (PAP) was combined with immunofluorescence for simultaneous localization of glutamate decarboxylase (GAD)-and somatostatin-like immunoreactivity in rat hippocampal neurons growing in dissociated cell culture. A subpopulation of GAD-immunoreactive neurons additionally exhibited somatostatin-like immunostaining. GAD-negative somatostatin-positive cells could not be observed. It is discussed whether the cultured somatostatin-containing GAD neurons correspond to a certain subclass of basket cells as they occur in situ.
Animals↗
Pathways of lymphocyte migration within the periarterial lymphoid sheath of rat spleen.
Male Wistar rats were injected intravenously with 5-(3H)uridine-labeled lymphocytes isolated from lymph nodes of syngeneic donors and enriched in T cells. After short periods of time (3 to 120 min after injection), labeled lymphocytes were localized in spleen compartments using autoradiography to identify routes of lymphocyte movement from blood into splenic parenchyma and to follow migration pathways of recirculating lymphocytes within the periarterial lymphoid sheath (PALS). Topographical analysis of labeled lymphocytes was performed in specific planes of PALS characterized by the diameter of the arterial vessel and termed PALS large, PALS medium, and PALS small (PALS L, PALS M, PALS S, respectively). Attention was also paid to accumulations of labeled lymphocytes close to the arterial vessel wall. Initially, labeled lymphocytes were localized in PALS S and PALS M near the terminal branching of arterial vessels and in the marginal zone (MZ). We conclude that lymphocytes emigrate from blood into splenic parenchyma within two white pulp compartments: in MZ, and directly within PALS through the wall of capillary vessels. The sequential accumulation of labeled cells near arterial vessels of increasing diameter suggests that the recirculating pool of lymphocytes migrates into the central part of PALS L by two routes: from MZ, and along arterial vessels from PALS S and PALS M.
Animals↗
Angiotensin II promotes development of neurophysin neurons in dissociated culture.
Dissociated cultures of retrochiasmatic hypothalamus of 18-day-old rat embryos were continuously treated with 2 X 10(-9) M angiotensin II (ang II) from the third day in vitro on. Cultivation was terminated at days 9 and 16 in vitro, respectively. Neurons immunoreactive for neurophysin, arg-vasopressin and oxytocin were visualized by immunocytochemistry, using the unlabeled antibody technique, and counted. Large, well-differentiated magnocellular neurons and small, probably immature cells could be distinguished. A certain number of large neurons were, in addition, immunoreactive for either vasopressin or oxytocin whereas the small cells were devoid of such staining. Ang II treatment brought about a modest increase in neurophysin-immunoreactive (NEU-IR) cell numbers at day 9 and a drastic augmentation of both large and small NEU-IR cells at day 16 in vitro, without appreciably affecting the total counts of neurons per culture.
Angiotensin II↗