Correlation between tyrosine hydroxylase activity and catecholamine concentration or turnover in brain regions.

Tyrosine hydroxylase activity correlated significantly with norepinephrine concentration and turnover, when results from regions containing predominantly noradrenergic terminals were compared, and with dopamine concentration and turnover when results from regions containing predominantly dopaminergic terminals were compared. Regions containing dopamine or norepinephrine cell bodies were characterized by higher tyrosine hydroxylase activities as compared to regions containing mostly nerve terminals. Higher levels of tyrosine hydroxylase activity and transmitter turnover were observed in regions containing dopaminergic terminals than in regions containing norepinephrine terminals. These findings are consistent with the view that tyrosine hydroxylase activity is linked to rates of catecholamine utilization by neurons in the CNS.     

Monosynaptic GABAergic signaling from dentate to CA3 with a pharmacological and physiological profile typical of mossy fiber synapses

Mossy fibers are the sole excitatory projection from dentate gyrus granule cells to the hippocampus, where they release glutamate, dynorphin, and zinc. In addition, mossy fiber terminals show intense immunoreactivity for the inhibitory neurotransmitter GABA. Fast inhibitory transmission at mossy fiber synapses, however, has not previously been reported. Here, we show that electrical or chemical stimuli that recruit dentate granule cells elicit monosynaptic GABA(A) receptor-mediated synaptic signals in CA3 pyramidal neurons. These inhibitory signals satisfy the criteria that distinguish mossy fiber-CA3 synapses: high sensitivity to metabotropic glutamate receptor agonists, facilitation during repetitive stimulation, and NMDA receptor-independent long-term potentiation. GABAergic transmission from the dentate gyrus to CA3 has major implications not only for information flow into the hippocampus but also for developmental and pathological processes involving the hippocampus.     

Reduced MPTP toxicity in noradrenaline transporter knockout mice

The noradrenergic neurons of the locus coeruleus (LC) are damaged in Parkinson's disease (PD). Neurotoxin ablation of the LC noradrenergic neurons has been shown to exacerbate the dopaminergic toxicity of MPTP, suggesting that the noradrenergic system protects dopamine neurons. We utilized mice that exhibit elevated synaptic noradrenaline (NA) by genetically deleting the noradrenaline transporter (NET), a key regulator of the noradrenergic system (NET KO mice). NET KO and wild-type littermates were administered MPTP and striatal dopamine terminal integrity was assessed by HPLC of monoamines, immmunoblotting for dopaminergic markers and tyrosine hydroxylase (TH) immunohistochemistry. MPTP significantly reduced striatal dopamine in wild-type mice, but not in the NET KO mice. To confirm that the protection observed in the NET KO mice was due to the lack of NET, we treated wild-type mice with the specific NET inhibitor, nisoxetine, and then challenged them with MPTP. Nisoxetine conferred protection to the dopaminergic system. These data indicate that NA can modulate MPTP toxicity and suggest that manipulation of the noradrenergic system may have therapeutic value in PD.     

Age-related decline in the tyrosine hydroxylase-immunoreactive innervation of the amygdala and dentate gyrus in mice

Numbers of catecholaminergic neurons are known to decline with aging. Whether projections of these neurons to the forebrain are similarly affected is not known. High densities of tyrosine hydroxylase-immunoreactive (TH-ir) fibers are found in the hippocampal formation (CA1-3, dentate gyrus) and in the amygdala of normal adult mice. We report here that densities of TH-ir fibers in the amygdala and hippocampus in aged mice (21-26 months) decrease dramatically and in a subregion-specific fashion. There is a reduction of 35% in the dentate gyrus, while hippocampal regions CA1 through CA3 are almost entirely spared. In the amygdala the lateral, basolateral, basomedial, and central nucleus were affected, with fiber reduction ranging from 19% to 34%. These results indicate that the age-related decline of TH-ir catecholaminergic cell bodies in the substantia nigra and the ventral tegmental area induces substantial losses of TH-ir fibers in the amygdala and dentate gyrus, but not in other areas of the hippocampal formation. This suggests that region-specific factors may be implicated in the regulation of maintenance vs. degeneration of TH-ir fibers during aging.     

Association of dopaminergic fibers with corticotropin releasing hormone (CRH)-synthesizing neurons in the paraventricular nucleus of the rat hypothalamus

Summary Catecholamines are known to exert a central influence on the hypothalamo-hypophyseal-adrenal neuroendocrine system. The selective dopaminergic innervation of the hypothalamic paraventricular nucleus (PVN) and putative relationships between dopaminergic fibers and corticotropin releasing hormone (CRH)-synthesizing neurons were studied in the male rat by means of immunocytochemistry following the elimination of noradrenergic and adrenergic inputs to the hypothalamus. A 3.0-mm-wide coronal cut was placed unilaterally in the brain at the rostral level of the mesencephalon. All neuronal structures from the cortex to the ventral surface of the brainstem, including the ascending catecholaminergic fiber bundles were transected. This surgical intervention resulted in the accumulation of dopamine--hydroxylase (DBH)-immunoreactivity in axons proximal to the cut, and an almost complete disappearance of DBH activity in those located distal to the lesion. Two weeks following the operation, DBH immunoreactivity was significantly diminished in the PVN located on the side of lesion, while tyrosine hydroxylase (TH)-immunoreactivity was present in a substantial number of fibers in the same nucleus. Both DBH- and TH-immunoreactive axons were preserved in the contralateral PVN. Simultaneous immunocytochemical localization of either DBH- or TH-IR fibers and corticotropin releasing hormone-synthesizing neurons in the hypothalami from brainstem-lesioned, colchicine treated animals revealed that the distribution of catecholaminergic fibers and CRH neurons is homologous within the PVN of the intact side. Only a few scattered DBH-immunoreactive axons were detected among CRH-producing neurons in the PVN on the side of the lesion. In contrast, many tyrosine hydroxylase containing neurons and neuronal processes were observed on the lesioned side and the TH-IR fibers established juxtapositions with CRH-synthesizing neurons.These morphological data demonstrate that following the surgical ablation of noradrenergic and adrenergic afferents to the PVN, a substantial number of tyrosine hydroxylase-IR fibers remained in the nucleus and they were associated with corticotropin releasing hormone synthesizing neurons. Therefore, it is hypothesized that the paraventricular nucleus receives a selective dopaminergic innervation and these dopaminergic axons might influence the function of the pituitary and adrenal glands via the hypothalamic CRH system.Supported by grants from the National Science Foundation (NSF INT 8703030), the Hungarian Academy of Sciences (OTKA 104), the National Institutes of Health (NS 19266) and the National Foundation of Technical Development (OKKFT Tt 286/1986)     

Association of dopaminergic fibers with corticotropin releasing hormone (CRH)-synthesizing neurons in the paraventricular nucleus of the rat hypothalamus

Catecholamines are known to exert a central influence on the hypothalamo-hypophyseal-adrenal neuroendocrine system. The selective dopaminergic innervation of the hypothalamic paraventricular nucleus (PVN) and putative relationships between dopaminergic fibers and corticotropin releasing hormone (CRH)-synthesizing neurons were studied in the male rat by means of immunocytochemistry following the elimination of noradrenergic and adrenergic inputs to the hypothalamus. A 3.0-mm-wide coronal cut was placed unilaterally in the brain at the rostral level of the mesencephalon. All neuronal structures from the cortex to the ventral surface of the brainstem, including the ascending catecholaminergic fiber bundles were transected. This surgical intervention resulted in the accumulation of dopamine-beta-hydroxylase (DBH)-immunoreactivity in axons proximal to the cut, and an almost complete disappearance of DBH activity in those located distal to the lesion. Two weeks following the operation, DBH immunoreactivity was significantly diminished in the PVN located on the side of lesion, while tyrosine hydroxylase (TH)-immunoreactivity was present in a substantial number of fibers in the same nucleus. Both DBH- and TH-immunoreactive axons were preserved in the contralateral PVN. Simultaneous immunocytochemical localization of either DBH- or TH-IR fibers and corticotropin releasing hormone-synthesizing neurons in the hypothalami from brainstem-lesioned, colchicine treated animals revealed that the distribution of catecholaminergic fibers and CRH neurons is homologous within the PVN of the intact side. Only a few scattered DBH-immunoreactive axons were detected among CRH-producing neurons in the PVN on the side of the lesion. In contrast, many tyrosine hydroxylase containing neurons and neuronal processes were observed on the lesioned side and the TH-IR fibers established juxtapositions with CRH-synthesizing neurons.(ABSTRACT TRUNCATED AT 250 WORDS)     

Spatiotemporal expression of sonic hedgehog signalling molecules in the embryonic mesencephalic dopaminergic neurons

Midbrain dopaminergic neurons (mDA) play an important role in controlling the voluntary motor movement, reward, and emotion-based behaviour. Differentiation of mDA neurons from progenitors depends on several secreted proteins, such as sonic hedgehog (SHH). The present study attempted to elucidate the possible role(s) of some SHH signaling components (Ptch1, Gli1, Gli2 and Gli3) in the spatiotemporal development of mDA neurons along the rostrocaudal axis of the midbrain and their possible roles in differentiation and survival of mDA neurons and the significance of using in vitro models for studying the development of mDA neurons. At E12 and E14, only Ptch1 and Gli1 were expressed in ventrolateral midbrain domains. All examined SHH signalling molecules were not detected in mDA area. Whereas, in MN9D cells, many SHH signalling molecules were expressed and co-localized with the dopaminergic marker; tyrosine hydroxylase (TH), and their expression were upregulated with SHH treatment of the MN9D cells. These results suggest that mDA neurons differentiation and survival might be independent of SHH in the late developmental stages (E12-18). Besides, MN9D cell line is not the ideal in vitro model for investigating the differentiation of mDA and hence, the ventral midbrain primary culture might be favored over MN9D line.     

Optogenetic Measurement of Presynaptic Calcium Transients Using Conditional Genetically Encoded Calcium Indicator Expression in Dopaminergic Neurons

Calcium triggers dopamine release from presynaptic terminals of midbrain dopaminergic (mDA) neurons in the striatum. However, calcium transients within mDA axons and axon terminals are difficult to study and little is known about how they are regulated. Here we use a newly-developed method to measure presynaptic calcium transients (PreCaTs) in axons and terminals of mDA neurons with a genetically encoded calcium indicator (GECI) GCaMP3 expressed in transgenic mice. Using a photomultiplier tube-based system, we measured electrical stimulation-induced PreCaTs of mDA neurons in dorsolateral striatum slices from these mice. Single-pulse stimulation produced a transient increase in fluorescence that was completely blocked by a combination of N- and P/Q-type calcium channel blockers. DA and cholinergic, but not serotoninergic, signaling pathways modulated the PreCaTs in mDA fibers. These findings reveal heretofore unexplored dynamic modulation of presynaptic calcium in nigrostriatal terminals.     

Tyrosine Hydroxylase Content of Residual Striatal Dopamine Nerve Terminals Following 6-Hydroxydopamine Administration: A Flow Cytometric Study

Fluorescence-activated cell sorting based on immunolabeling with a monoclonal antibody to tyrosine hydroxylase and a fluorescein-conjugated secondary antibody was used to identify striatal synaptosomes derived from nigrostriatal dopamine nerve terminals. The amount of tyrosine hydroxylase immunoreactivity in dopaminergic striatal synaptosomes prepared from control rats was compared to the amount in dopaminergic synaptosomes prepared from rats that had received intraventricular injections of 6-hydroxydopamine. Although the absolute number of dopaminergic synaptosomes was decreased in lesioned animals, those residual dopamine terminals present contained more tyrosine hydroxylase than did dopamine terminals from control rats. Both the decrease in the absolute number of dopamine terminals and the increase in tyrosine hydroxylase immunoreactivity in residual terminals were proportional to the extent of the lesion, as determined by measurement of striatal dopamine levels. These results suggest that an increase in the amount of tyrosine hydroxylase protein in residual terminals may represent one compensatory mechanism by which residual dopamine neurons maintain normal striatal function after partial destruction of the nigrostriatal dopamine projection.     

Dopamine transporter expression distinguishes dopaminergic neurons from other catecholaminergic neurons in the developing zebrafish embryo

To characterize the formation of the dopaminergic system in the developing zebrafish CNS, we cloned cDNAs encoding tyrosine hydroxylase (th), an enzyme in dopamine synthesis, and the dopamine transporter (dat), a membrane transport protein which terminates dopamine action by re-uptake. Dopaminergic neurons are first detected between 18 and 19 h post-fertilization in a cluster of cells in the ventral diencephalon. Subsequently, th and dat detection identifies dopaminergic neurons in the olfactory bulb, the pretectum, the retina and the locus coeruleus. Neurons expressing th but not dat are adrenergic or noradrenergic, and are found in the locus coeruleus, the medulla, the likely analog of the carotid body, and precursors of the enteric and sympathetic nervous system.     

Abnormal compartmentalization of norepinephrine in mouse dentate gyrus in alpha-synuclein knockout and A30P transgenic mice

In the dentate gyrus of the mouse hippocampus, presynaptic recruitment of norepinephrine in response to repeated-burst stimulation can be described in terms of an interaction between storage and readily releasable pools. The dynamics of this interaction depends on neuronal activity (bursting), so that the higher the demand for norepinephrine, the faster it is delivered from the storage pool. We also found that alpha-synuclein, a presynaptic protein that plays a crucial role in dopamine compartmentalization in the striatum, is also involved in the compartmentalization of norepinephrine in the dentate gyrus. Experiments in transgenic mice with modified or absent alpha-synuclein revealed that the familial Parkinson's disease-linked alpha-synuclein mutation A30P can cause selective changes in the function of noradrenergic terminals. Addition of mutated human alpha-synuclein abolished the normal norepinephrine mobilization. There were no compensatory mechanisms available in the norepinephrine presynaptic terminals. In contrast, deletion of mouse alpha-synuclein is compensated for by increased vesicle transport from the storage pool. The effects are essentially the same as previously reported for dopaminergic terminals in the striatum, indicating that the important role of alpha-synuclein in neurotransmitter mobilization is not limited to dopaminergic terminals.     

Cholinergic and catecholaminergic neurons relay striatal information to the optic tectum in amphibians.

In the amphibians Rana perezi and Xenopus laevis, the involvement of cholinergic and catecholaminergic neurons in the relay of basal ganglia inputs to the tectum was investigated. Tract-tracing experiments, in which anterograde tracers were applied to the basal ganglia and retrograde tracers to the optic tectum, were combined with immunohistochemistry for choline acetyltransferase and tyrosine hydroxylase. The results of these experiments suggest that dopaminergic neurons of the suprachiasmatic nucleus and pretectal region, noradrenergic cells of the locus coeruleus and the cholinergic neurons of the pedunculopontine and laterodorsal tegmental nuclei mediate at least part of the basal ganglia input to the tectum in anurans.     

Targeting the Hippocampal Mossy Fiber Synapse for the Treatment of Psychiatric Disorders

It is widely known that new neurons are continuously generated in the dentate gyrus of the hippocampus in the adult mammalian brain. This neurogenesis has been implicated in depression and antidepressant treatments. Recent evidence also suggests that the dentate gyrus is involved in the neuropathology and pathophysiology of schizophrenia and other related psychiatric disorders. Especially, abnormal neuronal development in the dentate gyrus may be a plausible risk factor for the diseases. The synapse made by the mossy fiber, the output fiber of the dentate gyrus, plays a critical role in regulating neuronal activity in its target CA3 area. The mossy fiber synapse is characterized by remarkable activity-dependent short-term synaptic plasticity that is established during the postnatal development and is supposed to be central to the functional role of the mossy fiber. Any defects, including developmental abnormalities, in the dentate gyrus and drugs acting on the dentate gyrus can modulate the mossy fiber-CA3 synaptic transmission, which may eventually affect hippocampal functions. In this paper, I review recent evidence for involvement of the dentate gyrus and mossy fiber synapse in psychiatric disorders and discuss potential importance of drugs targeting the mossy fiber synapse either directly or indirectly in the therapeutic treatments of psychiatric disorders.