Åke Ljungdahl

Distribution of substance P-like immunoreactivity in the central nervous system of the rat--I. Cell bodies and nerve terminals.

Abstract The distribution of substance P-immunoreactive structures in the central nervous system of young (1–3 weeks old), adult and colchicine-treated adult male rats has been studied, using the indirect immunofluorescence technique of Coons and collaborators. Substance P-positive cell bodies were observed in more than thirty areas including the spinal cord and many parts of the brain stem. Extensive networks of substance P-positive nerve terminals of varying densities were found in most areas of the central nervous system. The cerebral and cerebellar cortices contained only few substance P-positive structures. It was difficult to identify substance-P immunoreactive axons in the rats studied. Some pathways could, however, be described but further experimental studies are necessary to elucidate the projections of the substance P-immunoreactive neurons in the rat central nervous system.

Dopamine Nerve Terminals in the Rat Limbic Cortex: Aspects of the Dopamine Hypothesis of Schizophrenia

The existence of cortical dopamine nerve terminals is demonstrated with a highly sensitive modification of the Falck-Hillarp fluorescence technique. This confirms previous biochemical reports of high dopamine levels in the cortex. The histochemistry reveals that the distribution is regional and confined to the limbic cortex.

Studies on central and peripheral noradrenaline neurons using a new dopamine-β-hydroxylase inhibitor

Abstract The effects of a new dopamine-β-hydroxylase inhibitor, FLA63, have been studied on central and peripheral noradrenaline (NA) neurons using histochemical and biochemical analysis of monoamines. FLA63 caused a rapid and selective depletion of central NA stores without affecting dopamine (DA) and 5-hydroxytryptamine (5-HT) stores. The histochemical findings indicated that practically all the NA cell bodies in the lower brain stem were localized to the medulla oblongata and pons whereas the DA cell bodies were localized to the mesencephalon. FLA63 induced NA depletion was nerve impulse dependent. NA formation from dopa was markedly blocked by FLA63 treatment. The results with dopa suggest that DA cannot accumulate in high amounts in the NA storage granules. The in vitro studies showed neither NA uptake blocking activity nor any inhibitory effects on monoamine oxidase activity by FLA63. Studies on cold stress and FLA63 indicated that central NA neurons were not of critical importance in thermoregulation.

Pharmaco-histochemical evidence of the existence of dopamine nerve terminals in the limbic cortex

With a recent modification of the Falck-Hillarp technique using glyoxylic acid perfusion and Vibratome sectioning in combination with pharmalogical models for the selective demonstration of dopamine stores, evidence has been obtained that there exist large numbers of previously unknown systems of dopamine nerve terminals in the limbic cortex, e.g. in the gyrus cinguli, the entorhinal cortex and the amygdaloid cortex. No evidence for the existence of dopamine cell bodies in these areas was obtained.

Demonstration of central γ-aminobutyrate-containing nerve terminals by means of antibodies against glutamate decar☐ylase

Abstract A new procedure is described to obtain specific antibodies against mouse glutamate decar☐ylase using a purified glutamate decar☐ylase preparation obtained by affinity chromatography. Antibodies prepared against the Sepharose-bound glutamate decar☐ylase caused a concentration-dependent inhibition of glutamate decar☐ylase activity. Furthermore, the glutamate decar☐ylase antibodies gave a single precipitation band in double immunodiffusion, immunoelectrophoresis and in counterimmunoelectrophoresis analysis when reacted against crude extracts from mouse or rat brain. Immunohistochemical analysis in the rat brain demonstrated specific glutamate decar☐ylase-related immunofluorescence, presumably in γ-aminobutyrate-containing nerve terminals, but in no case in cell bodies in any part of the brain and the spinal cord. Two principal types of presumed γ-aminobutyrate-containing nerve terminals could be demonstrated. One type is strongly glutamate decar☐ylase immunoreactive, appears to have fairly large varicosities, may possibly make axosomatic and axodendritic contacts, and is confined mainly to the deep cerebellar nuclei, the nucleus vestibularis lateralis, the substantia nigra, the globus pallidus and the substantia innominata together with adjacent parts of the medial forebrain bundle and the dorsal part of the olfactory tubercle. In all these areas they are found in high densities. It is suggested, partly in agreement with previous findings that this type of γ-aminobutyrate-containing nerve terminal belongs to Golgi type I nerve cells (with long projections), contains high amounts of glutamate decar☐ylase enzyme protein and may mediate postsynaptic inhibition. The other type of γ-aminobutyrate-containing nerve terminal is weakly to moderately glutamate decar☐ylase immunoreactive, appears to have fine varicosities, may possibly make both axosomatic, axodendritic and axoaxonic contacts and is found all over the brain and the spinal cord in low to high densities. The distribution of this type correlates fairly well with the known distribution of glutamate decar☐ylase activity in the rat brain. It is suggested that this type of γ-aminobutyrate-containing nerve terminal mainly belongs to Golgi type II neurons (interneurons), contains relatively low amounts of glutamate decar☐ylase enzyme protein, and mediates both pre- and postsynaptic inhibition. The findings underline the existence of a new type of γ-aminobutyrate-containing Golgi type I neuron, controlling activity in the substantia innominata and in the outflow from Callejas islands of the olfactory tubercle and the view that γ-aminobutyrate may play a role in synaptic processes in all parts of the brain and the spinal cord.

Modification of the Falck-Hillarp formaldehyde fluorescence method using the vibratome®: simple, rapid and sensitive localization of catecholamines in sections of unfixed or formalin fixed brain tissue

SummaryTwo modifications of the original Falck-Hillarp formaldehyde fluorescence technique are presented, both based on a recently introduced instrument, the Vibratome®, which permits cutting of unembedded tissue with a section thickness down to 10 μ,.The first modification involves sectioning of unfixed tissue at a temperature below +5°C, subsequent air drying and reaction with formaldehyde vapours. In the second procedure formalin fixed tissue is cut and processed as described above. It is essential that both formalin fixation and cutting of the fixed tissue takes place at a low temperature to avoid diffusion of the catecholamines.The results show that with both techniques central CA neurons can be visualized with a high degree of sensitivity. Furthermore, since the sections are free from fractures—a common problem in freeze-dried tissues—the Vibratome® technique represents a valuable tool for mapping studies. It may also be added that since many steps of the original procedure are omitted the present techniques are also more rapid and simple. It is pointed out that using the Vibratome® procedure on formalin fixed tissue, it will be possible to combine e.g. cholinesterase staining or Fink-Heimer silver impregnation or immunofluorescent studies with the Falck-Hillarp technique on serial or even on the same sections.

CD8 is critically involved in lymphocyte activation by a T. brucei brucei-released molecule

T. brucei brucei released a lymphocyte triggering factor (TLTF), which triggered purified CD8+, but not CD4+, T cells to interferon gamma (IFN-gamma) mRNA expression and secretion and to [3H]thymidine incorporation. TLTF also induced mRNA for transforming growth factor beta, but not for interleukin-4. The action of this TLTF on mononuclear cell (MNC) cultures was blocked by anti-CD8 antibodies and by soluble CD8. MNCs from a mutant mouse strain lacking CD8 expression were not triggered by TLTF. IFN-gamma provides a growth stimulus for T. brucei brucei, and infected CD8- mice had much lower parasitemia and survived longer than CD8+ mice. The host-parasite interaction in experimental African trypanosomiasis thus involves parasite release of TLTF, which by binding to CD8 triggers CD8+ cells to produce the parasite growth-promoting cytokine IFN-gamma.

Memantine abrogates neurological deficits, but not CNS inflammation, in Lewis rat experimental autoimmune encephalomyelitis

Memantine, a clinically employed drug with N-methyl-D-aspartate (NMDA) receptor antagonistic effects, dose-dependently ameliorates neurological deficits in Lewis rat experimental autoimmune encephalomyelitis (EAE). Interestingly, this therapeutic effect was not due to dampened CNS inflammation, as assessed by immunohistochemical evaluation of spinal cord tissue. Furthermore, numbers of interferon gamma (IFN gamma) mRNA expressing cells were not decreased, as assessed by in situ hybridization. Systemic immunity in terms of numbers of IFN gamma secreting cells in response to immunodominant myelin basic protein (MBP) peptides ex vivo was not reduced, and non-toxic doses of memantine did not affect lymphocyte proliferation or IFN gamma secretion in vitro. Considering these findings, we hypothesize that effector mechanisms responsible for reversible neurological deficits in EAE may involve NMDA receptors, and this highlights neurons as targets during autoimmune neuroinflammation.

Immunohistochemical identification of two types of dopamine neuron in the rat olfactory bulb as seen by serial sectioning.

SummarySeveral neurons around the glomeruli in the rat olfactory bulb contain the enzyme tyrosine hydroxylase as revealed by light and electron microscopic immunohistochemistry. Electron microscopic analysis of serial sections revealed that both superficial tufted cells and small periglomerular neurons were labelled. These results give further support for the view that dopamine neurons in the rat olfactory bulb, from a neuroanatomical point of view, do not represent a homogeneous cell population. Furthermore, taken together with previous results in the literature our findings indicate that, from a transmitter histochemical point of view, neither tufted cells nor periglomerular neurons represent a homogeneous cell population.

Facial nerve transection causes expansion of myelin autoreactive T cells in regional lymph nodes and T cell homing to the facial nucleus

Nervous tissue expression of immunological signal and recognition molecules, as well as lymphoid tissue immune responses after facial nerve trauma was studied in male rats of the Lewis and Brown Norway (BN) strains. In both rat strains nerve transection caused within four days the appearance of IFN-gamma-like immunoreactivity in the cytoplasm of axotomized motor neurons and an induction of MHC class I and II, and CD4 molecules on surrounding glial cells to a similar extent. T lymphocytes also infiltrated the facial nuclei ipsilateral to the axotomy in all animals. The number of autoreactive T cells in superficial cervical lymph nodes, which in response to whole myelin or peptides of myelin basic protein (MBP) secreted IFN-gamma increased markedly after axotomy. This response was more conspicuous in Lewis rats, which are susceptible to experimental allergic encephalomyelitis (EAE), than in BN rats, which are EAE resistant. A proportion of the axotomized Lewis rats also developed widespread perivascular infiltration of mononuclear cells in the CNS, reminiscent of EAE. Hypothetically, a strong expansion of myelin autoreactive IFN-gamma producing T cells secondary to nerve trauma may have immunopathological consequences in genetically predisposed individuals. It is also possible that myelin reactive T cells, whether recruited to the lesioned nerve, could have impact on macrophage function during Wallerian degeneration in the distal stump.