Biomedical subjects
Publications and source records attributed to C Köhler.
Infusion of 120 nmol quinolinic acid into several regions of the rat's brain revealed differences in vulnerability to its neurotoxic effects, as judged by light microscopical analysis. The striatum, the pallidal formation and the hippocampus were the most susceptive brain areas whereas the cerebellum, substantia nigra, amygdala, medial septum and hypothalamus proved more resistant.
Using a specific antibody to the catecholamine (CA) synthesizing enzyme, tyrosine hydroxylase (TH), in combination with the avidin-biotin-peroxidase complex method, we have found evidence for the existence of a new CA-containing cell group extending from the orbitofrontal cortex through the olfactory and pyriform cortices in the brain of two species of monkey. The TH-positive perikarya, which are 4000-5000 in number, are situated within the outer layers of these cortices and also within the olfactory tubercle and horizontal limbs of the diagonal band of Broca. They have small (10-20 microns) somata of round or oval shape. A majority are bipolar with long, slender dendrites but some are small, multipolar with widely branching dendrites. The shape and laminar distribution of these TH-positive neurons suggest that they may serve functions as cortical interneurons.
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The retrohippocampal region of the rat brain was analyzed by using immunohistochemistry with specific antibodies against somatostatin (SOM) and vasoactive intestinal polypeptide (VIP). Specifically immunoreactive neurons and terminal processes were labeled with either the anti-SOM or anti-VIP antiserum and they were referred to as SOM-like immunoreactive (SOM-LI) or VIP-like immunoreactive (VIP-LI) neurons and processes, respectively. The retrohippocampal region was rich in neuronal cell bodies and terminal processes showing immunoreactivity for SOM and VIP. In the entorhinal area SOM-LI neurons were located mainly in layers IV through VI and the VIP-LI neurons were found mainly in layers I through III. Thick (70-120 microns) sections treated with the immunoperoxidase method to achieve a Golgi-like staining pattern showed that cytological differences existed between SOM- and VIP-positive neurons. SOM-LI neurons were usually multipolar, fusiform, or occasionally pyramidal while VIP-LI neurons were usually bipolar, stellate, or fusiform. SOM-LI and VIP-LI axons and preterminal processes were differentially distributed within the laminae of the retrohippocampal region. VIP-LI terminals were found throughout all layers except layer I. SOM-LI terminals were found primarily in the molecular layers of all areas, layer IV of the medical and lateral entorhinal areas, and in the angular bundle. Thus, SOM-LI and VIP-LI neurons are distinguished by their morphology and their different distribution within the cortical layers and areas of the retrohippocampal region.
The distribution of gamma aminobutyric acid (GABA)-containing neurons and nerve terminals was examined in the rat septal area by using specific antibodies to the enzyme glutamic acid decarboxylase (GAD) in combination with the avidin-biotin immunoperoxidase method. Whereas only a few GAD positive neurons were present in the septum of normal rats, the septal area of rats treated with colchicine, an inhibitor of fast axonal transport, showed numerous GAD-immunoreactive neurons. These neurons were evenly filled with GAD-immunoreactive material throughout the cytoplasm of the soma and proximal parts of the dendrites. Although GAD-positive neurons were present in most parts of the septal area, their density differed greatly in the different septal subnuclei. Both the diagonal band of Broca (vertical and horizontal parts) and the lateral septum were rich in GAD positive cell bodies, whereas the medial septal nucleus and the intermediate parts of the lateral septum contained relatively few. Within the lateral septum itself a larger number of labeled cell bodies was present in its ventral subdivision. The anterior hippocampal rudiment (taenia tecta) contained numerous GAD-positive neurons, while the septal component of the island of Calleja (insula magna) was devoid of them. GAD-immuno-positive neurons found within the septum ranged from small (15 microns) to large (30-35 micron). They were round or multipolar in the diagonal band, medium-sized multipolar in the lateral septum, and pyramidal, round or fusiform in the anterior hippocampal rudiment. GAD-immunoreactive nerve terminals are present in most subdivisions of the septal nuclei, with the exception of myelinated fiber tracts, and throughout all rostrocaudal levels of the septum. However, the density of the innervation is not the same within all individual nuclei. The lateral septum (dorsal and ventral parts) contained high density innervation but the diagonal band of Broca had a lower density of GAD-positive terminals. The lateral border of the islands of Calleja was rich in thick GAD-positive processes that appeared to be continuous with GAD-immunoreactive processes of the substantia inominata. The inner portion of the molecular layer adjacent to the granule cells of the anterior hippocampal rudiment contained a rich GAD-positive terminal field.
The retrograde axonal labeling of hippocampal GABA-ergic neurons was studied after in vivo injections of a characterized antibody against glutamic acid decarboxylase (GAD) into different parts of the hippocampal region. Small injections (50 nl) of undiluted GAD antibody into the area dentata (AD) labelled fusiform and dentate pyramidal basket cells within the AD and fusiform and multipolar cells in subfields CA2/CA3 (a and b) of Ammon's horn. The labeled cells were characterized by intense immunoreactivity of the soma and proximal parts of the dendrites, while the nucleus contained little or none. The morphological appearance and laminar positions of these cells corresponded to hippocampal GAD-positive neurons, as shown previously (Ribak et al. 1978) with immunocytochemistry. Injections of anti-GAD into the medial entorhinal area, subiculum, and CA1 region labeled cells in strata oriens, pyramidale and radiatum of CA2 and CA3a, but injections of the antibody into these latter areas failed to label cells in the medial CA1 and subiculum, thus suggesting a preferential organization of hippocampal GABA neuronal projections in a lateral to medial direction. Injections of preimmune sera or antiserum preabsorbed with the pure enzyme antigen GAD failed to label cells in a manner similar to that described for the anti-GAD injections. These observations, taken together with the finding that injections of anti-GAD into the terminal field of non-GABA-ergic pathways never resulted in retrograde axonal transport of the antibody-antigen complex, suggest that the in vivo injection of GAD antibody is a useful method to study the organization of hippocampal GABA-ergic neurons and their projections.
The neurotoxic properties of ibotenate and kainate after intracerebral application were compared in several regions of the rat brain. Ibotenate, being 5-10 times less toxic than kainate, caused lesions which were generally found to extend spherically from the tip of the injection cannula. In contrast, kainate injections often resulted in neuronal degeneration distant from the site of infusion, thus severely limiting its use as a tool for causing lesions in neurobiological studies. In some of the brain regions examined (hippocampus, septum), neurons appeared differentially susceptible to kainate but uniformly vulnerable to ibotenate. Some cell groups, such as those in the medial septum and the locus coeruleus, proved highly resistant to kainate but could be selectively ablated by ibotenate. These findings, together with differences between the two toxins in the evolution of neuronal degeneration (exemplified here in the hippocampal formation), appear to support previous suggestions that ibotenate and kainate exert their excitotoxic actions via different mechanisms. On the other hand, neuropathological changes caused in the cerebellum did not differ, since both ibotenate and kainate preferentially destroyed granule cells. Two nuclei, the arcuate nucleus of the hypothalamus and the nucleus of the fifth nerve, were found to be extremely resistant to either neurotoxin.
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Using antibodies to somatostatin in combination with the avidin-biotin immunohistochemical method, the laminar distribution and morphology of somatostatin-like (SOM-L) immunoreactive cells have been studied in the hippocampal formation of the rat. SOM-L positive cells are present throughout the longitudinal axis of the hippocampus. Most of these neurons are of medium (15-20 micrometers) to large (25-30 micrometers) size with soma of multipolar or fusiform shape. No pyramidal or granule cells contained SOM-L immunoreactivity. SOM-L-positive cells show a preferential distribution in the stratum oriens and stratum pyramidalae of Ammon's horn, and match the description of the polymorphic cells of the hilus area dentata. This geographic segregation may imply a chemical compartmentation of function in the hippocampus.
The distribution and morphology of vasoactive intestinal polypeptide-like (VIP-L) immunoreactive neurons were studied in thick (70-120 microns) sections from the rat hippocampal formation using VIP antibody in combination with immunohistochemistry. Analysis of serial sections cut through the regio superior revealed a relatively dense aggregation of cells in stratum lacunosum-moleculare. Classification of cells in these (and other) layers on the basis of their morphology and orientation showed that hippocampal VIP-positive cells, unlike cortical ones, constitute a remarkably heterogeneous population. Studies of axonal trajectories of individual VIP cells showed that while some have local terminal arbors within the stratum lacunosum-moleculare close to the parent soma, other VIP cells send longer projections to other layers and subfields of the hippocampal formation.
The distribution of cholecystokinin (CCK)-like (CCK-L) immunoreactive cells and nerve terminals was studied in the brains from rats and guinea pigs by using antibodies to the octapeptide cholecystokinin (CCK-8). Analysis of serial horizontal and sagittal sections through the retrohippocampal region in colcicine-pretreated rats revealed a relatively large number of CCK-L immunoreactive cells in the pre- and parasubiculum, subiculum, and the medial and lateral entorhinal area (EA) at all dorsal to ventral levels of the region. In the EA, the CCK-positive cells were scattered in all layers without any clear pattern. Analysis of CCK-positive cells in the retrohippocampal region showed that these cells form a morphologically heterogeneous group. The types of CCK-L immunoreactive cells ranged from small (approximately 10 micrometers) round, ovoid, or fusiform to large (approximately 30 micrometers) multipolar and pyramidal. CCK-L immunoreactive nerve fibers and preterminal processes were unevenly distributed in the retrohippocampal region. The densest innervation was found in the parasubiculum, subiculum, and the ventrolateral entorhinal area. Only a few scattered fibers were detected in the molecular layers of these structures and the outer layers of the presubiculum. Within the EA and CCK innervation indicated a heterogeneous laminar distribution that was densest in layers II and IV of the medial and lateral EA and diffuse in layers I and II. In layer II the immunoreactive nerve terminals encircled the pyramidal cell bodies, while in layers IV to VI and the most ventral part of lateral entorhinal area (LEA) and the transitional area between LEA and piriform cortex the CCK processes were distributed in a netlike fashion without clear relation to the cytoarchitectural characteristics of the area.
Large (200 nl) intraseptal injections of horseradish peroxidase (HRP) resulted in retrograde axonal labeling of both pyramidal and non-pyramidal neurons throughout all septo-temporal levels of the hippocampal formation in the rat brain. Small (50 nl) injections of HRP into the medial septum labeled cells of non-pyramidal shape in the stratum oriens and the stratum radiatum of regio inferior, stratum oriens of regio superior and the hilus of the area dentata. Small (50 nl) injections of HRP restricted to the lateral septum resulted in retrograde labeling of pyramidal cells in regio inferior and regio superior without labeling of non-pyramidal cells. These results suggest a new efferent projection system from the hippocampus consisting of non-pyramidal neurons which innervate the medial septum/diagonal band complex in the rat brain.
The distribution of 5-hydroxytryptamine (serotonin, 5-HT)-containing nerve fibers and terminals in the septal area of the rat was studied by using immunohistochemistry with specific antibodies to 5-HT in combination with fluorescent retrograde tracing methods. The 5-HT innervation of the septum is heterogeneous with regard to both the morphology of individual processes and the density of distribution in different parts of the septum. Three major classes of 5-HT like immunoreactive processes can be distinguished: 1) thin, convoluted fibers with small, round or elongated varicosities; 2) thick and relatively straight fibers with few varicosities; and 3) pericellular plexuses with large varicosities in close association with perikarya in the lateral septum. Three areas of the septum receive a prominent innervation by 5-HT processes: the diagonal band of Broca, the ventral part of the lateral septum, and an area bordering the medial edge of the islands of Calleja (insula magna). Whereas the two latter areas contain dense terminal networks, the diagonal band of Broca is occupied primarily by 5-HT fibers en route to other parts of the septum. Intraseptal injections of HRP or fluorescent dyes (granular blue, propidium iodide) resulted in retrograde labeling of neuronal cell bodies in several nuclei of the brainstem which are known to contain 5-HT neurons: the dorsal raphe, the median raphe, the nucleus reticularis tegmenti pontis, the raphe pontis, and the raphe magnus. Where fluorescent retrograde tracing was performed with 5-HT immunohistochemistry on the same tissue section, a prominent 5-HT containing pathway and a non-5-HT-containing pathway from the raphe nuclei to the septum were revealed. Finally, double retrograde fluorescent labeling after injections of granular blue or propidium iodide into the septum and entorhinal area respectively of the same rat revealed extensive branching of the raphe efferents. Thus, individual raphe neurons may simultaneously connect with septum and the entorhinal area, two structures essential for normal hippocampal function.
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The work presented here concerns the way in which the dorsal midbrain tegmentum (DMT) participates in the control of sexual behavior. It was first established that electrolytic DMT lesions accelerate mating in the male rat, primarily by abbreviating the post-ejaculatory interval. Since the effective lesions were accompanied by decreases in the in vitro synaptosomal uptake of [3H]noradrenaline (NA) in hippocampus and hypothalamus, the behavioral effects of DSP4 (which elicits degeneration in NA nerve terminals derived primarily from the locus coeruleus) were examined. The long-term behavioral consequences of DSP4, however, were to decrease copulatory rate despite substantial NA denervation of brain and spinal cord. Ibotenic acid-induced neuronal degeneration in the DMT, on the other hand, accelerated copulatory behavior while leaving NA innervation of hippocampus and hypothalamus unaffected. The magnitude of the behavioral effect in ibotenic acid-treated rats was less than that induced by electrolytic DMT lesions. It is tentatively suggested on the basis of these-experiments that DMT cell bodies may form part of a system regulating sexual arousal mechanisms, whilst activity in a non-adrenergic fiber system running in the dorsal tegmental bundle may be required for active inhibition of sexual behavior after ejaculation. In additional experiments it was found that DSP4 treatment of female rats produced negligible effects on sexual behavior, estrous cyclicity and processes related to lactation.
It is well known that electrolytic lesions in the medial preoptic area (MPOA) and the lateral hypothalamic area (LHA) seriously impair masculine sexual behavior in the rat. We here report that bilateral infusions of the neurotoxin, ibotenic acid (IBO), in the MPOA were as effective as electrolytic lesions in eliminating copulation whereas no behavioral effects were detected following similar infusions in the LHA. Histological examination of MPOA and LHA following IBO exposure revealed extensive degeneration of neuronal cell bodies with little evidence of non-specific damage. Also, immunohistochemical studies suggested that the serotonergic innervation of the MPOA remained largely intact in spite of IBO treatment; similarly, the damage inflicted by IBO in LHA on tyrosine hydroxylase-immunoreactive fibers in the medial forebrain bundle was insignificant. These data suggest that: (i) the functional integrity of MPOA nerve cell bodies is necessary for the expression of sexual behavior, and (ii) disruption of mating produced by electrolytic LHA lesions is due to disruption of medial forebrain bundle fiber systems. Behavioral observations of non-copulating males suggested that the MPOA injury did not interfere with all aspects of their sexual interaction with the estrous female; rather, they appeared specifically unable to perform the reflexive pelvic thrust pattern normally associated with mounting. We here report, however, that the ability to perform mounts with pelvic thrusts was temporarily restored in the vast majority of MPOA-injured males by the i.p. administration of the ergot derivative, lisuride. About 50% of these MPOA-damaged males even ejaculated, often after a low number of intromissions and short ejaculation latencies. On the other hand, injections of naloxone (an opiate receptor antagonist) failed to activate mounting in MPOA-lesioned or castrated rats. On the basis of these findings the possible ways in which steroid hormone-sensitive brain areas might interact with monoamine-containing pathways are discussed.U
We have studied the localization of serotonin- and non-serotonin-containing cell bodies in the midbrain raphe nuclei that project to the entorhinal area and the hippocampal formation in the rat brain, using the technique of combined retrograde fluorescent tracing and immunohistochemistry on the same tissue section. The branching properties of these neurons were studied by retrograde double labelling using two fluorochromes which emit fluorescence with different spectral characteristics. After injections of granular blue or propidium iodide into the medial entorhinal area, retrogradely-labelled cells were found situated bilaterally in the caudal half of the dorsal raphe nucleus, the medial part of the median raphe and throughout the rostrocaudal extension of the nucleus reticularis tegmentipontis. Injections placed successively more laterally in the entorhinal area labelled progressively less cells contralaterally in the dorsal raphe and the reticular tegmental nucleus of the pons. After fluorochrome injections into the dorsal part of the hippocampal formation, retrogradely-labelled cells were found in the caudal part of the dorsal raphe, in the peripheral part of the median raphe and to a minor extent in the medial part of this nucleus, but not in the nucleus reticularis tegmentipontis. The experiments with double retrograde fluorescent tracing showed that the raphe nuclei do not send bilateral projections to the entorhinal area in spite of the fact that many of these cells are located contralateral to the injected hemisphere in single labelling experiments. Injections of the fluorochromes into the entorhinal area and hippocampal formation showed that at least 10% of the raphe cells project to both areas simultaneously. Analysis of sections incubated with antiserum to serotonin showed that a majority of the retrogradely-labelled versus serotonin-immunoreactive cells was found to vary within different parts of the individual raphe nuclei: the ventromedial part of the dorsal, the medial part of the median and the nucleus reticularis tegmentipontis being the highest. The findings indicate that both serotonin- and non-serotonin-containing neurons in the raphe innervate the hippocampal region, that these projections may be crossed but not bilateral, and that the same neuron in the raphe may influence the neural activity in the entorhinal area and the hippocampus simultaneously.
Tail vein injections of [3H]N-propylnorapomorphine ([3H]NPA) in male mice resulted in a dose-related accumulaton of radioactivity in the following brain regions: striatum (max), olfactory tubercle and cerebellum (min). The specific binding was saturable with increasing concentrations of the drug and stereospecifically displaced by (+) butaclamol. Dopamine agonist (apomorphine, NPA and bromocriptine) and antagonists (spiperone, haloperidol, (+) butaclamol and I-sulpiride) all caused dose-dependent prevention of [3H]NPA binding. Mianserin, phenoxybenzamine and propranolol did not prevent the in vivo [3H]NPA binding suggesting that [3H]NPA binds specifically to dopamine receptors in the striatum and the olfactory tubercle of the mouse.