Homospecific activity (activity per enzyme protein) of tyrosine hydroxylase increases in parkinsonian brain

Summary Tyrosine hydroxylase (TH) contents in the caudate nucleus, putamen, and substantia nigra from control and parkinsonism brains were measured for the first time by a sandwich enzyme immunoassay. Both the TH protein content and TH activity (V_max) were decreased in parallel in the parkinsonian brains as compared with those of the control brains. In contrast, TH homospecific activity (activity per enzyme protein) was significantly increased in the parkinsonian brains. The results indicate that the decrease of TH activity in parkinsonian brains is due to the decrease of TH protein content as a result of cell death. The increase in the homospecific activity of residual TH in parkinsonian brain suggests such molecular changes in TH molecules as result in a compensatory increase in TH activity.     

Glutamine inhibits the accumulation and hydroxylation of tryptophan in rat striatal synaptosomes

l-Glutamine at the concentration present in cerebrospinal fluid decreases the steady-state accumulation of the aromatic amino acids tryptophan, tyrosine and dihydroxyphenylalanine (DOPA) in rat striatal synaptosomes. Glutamine significantly inhibits synaptosomal tryptophan hydroxylase activity; it has less marked effects on tyrosine hydroxylase and DOPA decarboxylase activities. Thus, interaction between glutamine and tryptophan transport into nerve terminals may be one of the factors regulating the rate of serotonin synthesis in vivo.     

Tyrosine hydroxylase phosphorylation in rat brain striatal synaptosomes

The present studies were carried out to determine if tyrosine hydroxylase phosphorylation in rat brain striatal synaptosomes is activated by dibutyryl cyclic AMP treatment. Incubation of synaptosomes with [32P]orthophosphate, followed by immunoprecipitation and sodium dodecyl sulfate polyacrylamide gel electrophoresis, produced a band of radioactivity associated with a 62 kDa polypeptide. Treatment with the catecholamine neurotoxin, 6-hydroxydopamine, produced parallel losses of: (1) tyrosine hydroxylase enzyme activity, (2) dopamine content, and (3) the 62 kDa band of radioactivity. These data support the identification of this band as a tyrosine hydroxylase-derived polypeptide. Incubation with dibutyryl cyclic AMP produced an increase in soluble tyrosine hydroxylase activity and phosphorylation. These results suggest that the increase in synaptosomal catecholamine synthesis produced by dibutyryl cyclic AMP is mediated by an increase in tyrosine hydroxylase phosphorylation.     

β-Phenylethylamine reversal of chlorpromazine-induced activation of striatal tyrosine hydroxylase and catalepsy

The ability of β-phenylethylamine (PEA) to reverse (1) chlorpromazine-induced activation of striatal tyrosine hydroxylase, and (2) catalepsy produced by chlorpromazine, was examined. PEA, in a dose of 75 mg/kg, caused approximately 50% reduction in the degree of tyrosine hydroxylase stimulation produced by 20 mg/kg chlorpromazine. After 150 mg/kg PEA, complete reversal of tyrosine hydroxylase activation and partial reversal of catalepsy was observed. In these experiments, PEA was found to be about 10 times less potent than amphetamine and 25 times less ppotent than apomorphine. Thus, the ability of PEA to reverse the neurochemical and behavioral effects of striatal dopamine blockade is similar to known dopamine agonists.     

Evidence that adenosine A2 and dopamine autoreceptors antagonistically regulate tyrosine hydroxylase activity in rat striatal synaptosomes

Incubation of rat striatal synaptosomes with the adenosine receptor agonist 2-chloroadenosine (2-CADO) produced a concentration-dependent increase of dopamine (DA) synthesis (about 50% of control value). The effect was not additive with the stimulation produced by either 10 microM forskolin or 2 mM dibutyryl cyclic AMP. Pretreatment of striatal synaptosomes with 2-CADO produced an activation of tyrosine hydroxylase (TH) which withstood washing and lysing of the tissue. This activation was largely independent of the presence of Ca2+ ion in the preincubation medium and, when analyzed as a function of different concentrations of the pterin cofactor 6-methyl-5,6,7,8-tetrahydropterin (0.08-0.4 mM), it was associated with an apparent increase in the Vmax of the enzyme. Quinpirole, a selective D2 DA receptor agonist, reduced control synaptosomal DA synthesis and caused a persistent inhibition of TH activity. When added together with 2-CADO, quinpirole depressed the stimulation of DA synthesis and TH activity produced by the adenosine analog. The effect of quinpirole was stereospecifically antagonized by the D2 DA antagonist sulpiride. Quinpirole also inhibited the activation of TH elicited by a submaximal concentration of forskolin, but not that produced by dibutyryl cyclic AMP. The inhibitory effect of quinpirole on basal and 2-CADO-stimulated TH activities was mimicked by DA. These results indicate that presynaptic DA autoreceptors and adenosine A2 receptors interact antagonistically in controlling DA synthesis in rat striatal synaptosomes presumably by exerting opposite inputs on a presynaptic adenylate cyclase system.     

Dopamine differentiation factors increase striatal dopaminergic function in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned mice

We have previously shown that muscle-derived differentiation factors (MDF) and human recombinant acidic fibroblast growth factor (aFGF) have beneficial behavioral and neurochemical effects on the nigrostriatal dopaminergic neurons of 6-hydroxy-dopamine (6-OHDA)-lesioned rats (Jin and Iacovitti: Neurobiol Dis 2:1–12, 1995). In the present study, we determined the effects of similar treatments on mice treated with the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Five days after unilateral striatal infusion of MDF or aFGF into MPTP-lesioned mice, striatal tyrosine hydroxylase (TH) activity and dihydroxyphenylacetic acid (DOPAC) levels were bilaterally increased (20–35%) compared to untreated (lesion only) or control (phosphate buffered saline + bovine serum albumin) mice. These increases, however, were not accompanied by change in dopamine (DA) levels, indicating an elevation of DA synthesis (TH/DA) and turnover (DOPAC/DA). The present findings that MDF and aFGF may have neurochemical effects in vivo on the lesioned nigrostriatal dopaminergic system suggest their potential pharmacological role in the treatment of Parkinson's disease. © 1996 Wiley-Liss, Inc.     

Quantitative immunocytochemistry of tyrosine hydroxylase in rat brain. I. Development of a computer assisted method using the peroxidase-antiperoxidase technique

We sought to develop a quantitative immunocytochemical procedure using the peroxidase-antiperoxidase (PAP) technique to analyze amounts of the neurotransmitter biosynthetic enzyme tyrosine hydroxylase (TH) within the nucleus locus coeruleus (LC) of rat brain. Rats were perfused with 4% paraformaldehyde. The brains were embedded in paraffin, sectioned at 5 μm in the sagittal plane and immunocytochemically stained with antibodies to bovine adrenal TH. Staining intensity, measured by a TV image analysis system was reproducible within±5%.Reaction conditions required so that the intensity of the PAP reaction product was directly and linearly related to the amount of TH enzyme protein in tissue were obtained by reacting tissues with saturating concentrations (2.5 mM) of diaminobenzidine (DAB) substrate, a constant dilution of antibody, and incubation time adjusted so that the darkest elements in tissue were below saturation.Variations in staining intensity of serial sections through the LC were found to be insignificant when compared to variations in staining between animals.In order to increase the amount of immunoreactive TH in the LC, groups of rats were treated with reserpine. Immunocytochemical and biochemical analyses were performed in parallel groups of control and reserpine animals allowed to survive 1–3 days following a single injection (10 mg/kg s.c.) of reserpine.A close correlation was found to exist between the amount of TH enzyme protein determined biochemically and the density of staining for TH. The maximal increase in TH measured immunocytochemically was 2.2-fold which was about 80% of the maximal induction determined biochemically. We conclude that the PAP method can be used for quantitative immunocytochemistry of brain TH providing that optimal reaction conditions are established.     

Effects of dexamethasone and other glucocorticoid steroids on tyrosine hydroxylase activity in the superior cervical ganglion

Dexamethasone is known to elicit an increase of tyrosine hydroxylase activity in the superior cervical ganglion. The details of such a glucocorticoid effect were investigated in the present study. Of 4 glucocorticoids (dexamethasone, corticosterone, hydrocortisone and triamcinolone) examined in rats, only the synthetic steroid dexamethasone was found to be effective in increasing ganglionic tyrosine hydroxylase activity (by 50% at 48 h after drug administration). Corticosterone even at doses as high as 50 mg/kg failed to show an effect. Since recent reports indicate that a cytoplasmic glucocorticoid receptor is not present in the sympathetic ganglion, it is unlikely that the dexamethasone effect involves a receptor-mediated mechanism. Moreover, the dexamethasone effect was totally blocked by chlorisondamine, a nicotinic cholinergic receptor antagonist. The possibility of an enhanced impulse flow from the CNS, however, was excluded by the finding that decentralization immediately prior to dexamethasone administration did not prevent the increase of ganglionic tyrosine hydroxylase activity, although earlier decentralization (24 h or longer) abolished the steroid effect. Significantly, in the freshly decentralized ganglia, the increase of tyrosine hydroxylase activity by dexamethasone was still blocked by chlorisondamine. Since synaptic activity in terminals is known to continue for a brief period following nerve transection, our data support the contention that the primary site of the dexamethasone effect may be the preganglionic terminals.     

Nerve growth factor-mediated stimulation of tyrosine hydroxylase activity in a clonal rat pheochromocytoma cell line

In order to confirm the multiple neurotransmitter biosynthetic ability, the possibility of separation of the activities of tyrosine hydroxylase (TH), choline acetyltransferase and glutamic acid decar☐ylase was tested by subcloning of a clonal rat pheochromocytoma PC12 cell line. All of 9 subclones obtained showed significant activities of above 3 enzymes, indicating that the PC12 cell has multi-functional properties of neurotransmitter syntheses.One of the subclones, designated PC12h, was demonstrated to have nerve growth factor- (NGF) responsive TH activity. The ED₅₀ value of NGF to increase the TH activity was 1.7 ng/ml (6.5 × 10⁻¹¹M). A simultaneous addition of saturating amounts of NGF (50 ng/ml) and dexamethasone (10⁻⁶ M) resulted in the increase of TH activity that is equal to the sum of those achieved when either effector was added separately, indicating that the NGF-mediated increase of TH activity in PC12h cells was independent upon the effect of dexamethasone. And also, the TH activity increased by NGF was somewhat potentiated in PC12h cells cultured in a hormone-supplemented serum-free medium.     

Alterations in tyrosine hydroxylase activity elicited by raphe nuclei lesions in the rat locus coeruleus: evidence for the involvement of serotonin afferents

The time course of the variations in tyrosine hydroxylase (TH) activity was measured in the rat locus coeruleus (LC) after lesions of the nucleus raphe dorsalis (NRD), nucleus raphe centralis superior (NRCS) and nucleus raphe pontis (NRP). A certain number of lesions were performed in the raphe magnus (RM), the caudal and rostral NRP and the caudal and rostral NRCS, lateral to raphe nuclei and in adrenalectomized animals. The serotonin (5-HT) content in the LC was also determined after these lesions. Only raphe nuclei producing significant decreases in the 5-HT content in the LC are successful in provoking increases in the TH activity in the LC, thus these results suggest that the noradrenaline (NA) synthesis in the LC may be regulated by 5-HT afferents. Moreover, intraventricular injections of 5,6-dihydroxytryptamine (5,6-DHT) and administration of parachlorophenylalanine (PCPA) also produce significant increases in TH in the LC. After immunotitrations of TH in the LC it was shown that, with exception of a high dose of 5,6-DHT (75 micrograms), all these treatments provoke an increase in the concentration of the enzyme. It therefore seems that one of the functional roles of 5-HT in the LC could be the regulation of the concentration of TH.     

Ontogeny of tyrosine hydroxylase and dopamine-beta-hydroxylase activity in discrete limbic and hypothalamic structures of female rats

Tyrosine hydroxylase (TOH) and dopamine-ß-hydroxylase (DBH) activities were measured in the mediocortical amygdala (AMY), hippocampus (HPC), nucleus accumbens (ACB), medial preoptic area (MPO), and anterior and posterior parts of the mediobasal hypothalamus (AMBH and PMBH) in female rats of various postnatal ages. Serum LH, FSH and prolactin levels were also measured. In a previous study it was shown that dopamine (DA) and norepinephrine (NE) turnover rates in the MPO and the MBH increase between days 20 and 30 after birth and decrease thereafter. The cause for increased DA turnover was shown to be increasing serum prolactin levels. The activity of TOH in the ACB, MPO, AMBH and PMBH did not parallel DA turnover rates. Around day 20 after birth, when serum prolactin levels and preoptic and hypothalamic DA turnover increase, TOH activity also increased in the ACB and PMBH. In adult, diestrous animals, however, which have low serum prolactin levels and low preoptic and hypothalamic DA turnover, TOH activity was higher than in immature rats. The activity of DBH in the MPO, AMBH, AMY and HPC was high in 15-day-old animals, decreased between days 20 and 30 and then increased again to adult values. This pattern is just opposite to the one observed for serum prolactin levels and for preoptic and hypothalamic NE turnover. It is concluded that activity of both enzymes is not a good measure for neuronal activity of those NE and DA cells which innervate limbic and hypothalamic structures.     

GDNF abates serum deprivation-induced tyrosine hydroxylase Ser19 phosphorylation and activity

High dopamine levels can contribute to neuronal dysfunction, impair plasticity and be toxic to neuronal cells in pathological conditions. The synthesis of dopamine is regulated by phosphorylation of the rate-limiting enzyme tyrosine hydroxylase (TH) under physiological conditions, with the phosphorylation of Ser31 and Ser40 directly increasing TH activity. Although a third phosphorylation site, Ser19, does not appear to directly regulate TH activity in physiological conditions, its role in pathological conditions is poorly understood. In this study, we examined the effects of serum deprivation (to mimic loss of retrogradely/anterogradely transported target-derived neurotrophic factors following axonal injury) and glutamate receptor stimulation (to mimic excitotoxicity) on TH phosphorylation and activity in a cell line and in mesencephalic primary culture cells. In addition, we also tested whether glial cell line-derived neurotrophic factor (GDNF) can alter these changes. We demonstrate that serum-deprivation resulted in a sustained increase in Ser19 phosphorylation beginning at 3 h and lasting up to 10 h without any detectable change in Ser31 or Ser40 phosphorylation within this time frame. This increase in Ser19 phosphorylation was associated with enhanced TH activity and was due, in part, to glutamate-receptor-mediated calcium influx and possibly calcium/calmodulin-dependent protein kinase II (CaMKII) activation. Interestingly in this serum-deprivation model, GDNF blocked the increase in Ser19 phosphorylation and TH activity at the 10-h time point following serum deprivation. Furthermore, GDNF also blocked the glutamate-mediated increase in Ser19 phosphorylation in rat primary mesencephalic neuronal cultures. Taken together, these findings suggest that GDNF may reduce dopamine synthesis in pathological conditions.     

Repeated immobilization stress alters tyrosine hydroxylase, corticotropin-releasing hormone and corticosteroid receptor messenger ribonucleic Acid levels in rat brain

In situ hybridization histochemistry was used to localize and quantify the effects of acute and repeated immobilization stress on mRNA levels of tyrosine hydroxylase (TH) in catecholaminergic neurons in the locus ceruleus and substantia nigra and on mRNA levels of relevant markers of the hypothalamic-pituitary-adrenal axis, namely corticotropin-releasing hormone (CRH) in the hypothalamic paraventricular nucleus (PVN), proopiomelanocortin in the pituitary, and mineralocorticoid receptors (MR, type I) and glucocorticoid receptors (GR, type II) in the hippocampus, PVN and pituitary. Control, acutely stressed (1 × lMO, sacrificed immediately after 2 h of immobilization), and repeatedly stressed (6 × IMO plus delay, sacrificed 24 h after 6 daily 2-h immobilizations and 6 × lMO plus challenge, sacrificed immediately after the seventh daily 2-h immobilization) male Sprague-Dawley rats were examined. TH mRNA expression was increased in the locus ceruleus in the acutely stressed and repeatedly stressed animals. The increase in TH mRNA levels was greatest in the repeatedly stressed (6 × IMO plus challenge) group. TH mRNA levels were not altered in the substantia nigra. CRH mRNA levels in the PVN were significantly increased in the three stressed groups and the increase was greatest in the 6 × IMO plus challenge group. CRH mRNA levels were increased in the central nucleus of the amygdala only after acute stress. Proopiomelanocortin mRNA levels were elevated in the anterior pituitary during acute and repeated stress, but the magnitude of the effect was largest after acute stress. The changes in the hypothalamic-pituitary-adrenal axis were accompanied by an acute stress-induced increase in MR mRNA levels in the hippocampus, MR and GR mRNA levels in the PVN and GR mRNA levels in the pituitary. MR mRNA levels continued to be elevated in the PVN in the 6 × IMO plus challenge animals. Plasma corticosterone levels were elevated in the acute and repeated stress conditions. The results show that repeated immobilization stress produces a rapid and persistent increase in mRNA expression of TH in the locus ceruleus, CRH in the PVN, and proopiomelanocortin in the anterior pituitary. The TH-containing neurons in the locus ceruleus and the CRH-containing neurons in the PVN appear to preserve the capability to respond to repeated stimulation (6 × IMO plus challenge) indicating altered feedback mechanisms under repeated stress conditions. GR and MR mRNA levels are differentially regulated in the hippocampus, PVN and pituitary by acute and repeated stress. It is of interest that the central nervous system systems which are activated during repeated stress, namely the locus ceruleus-norepinephrine system and hypothalamic-pituitary-adrenal axis, are dysregulated in melancholic depression. Further studies of the central nervous system effects of prolonged exposure to stress may help elucidate the mechanisms underlying dysregulation of the locus ceruleus-norepinephrine system and hypothalamic-pituitary-adrenal axis in depression and other stress-related psychiatric diseases.     

Mechanisms of MPTP (1-methyl-4-phenyl-l, 2, 3, 6-tetrahydropyridine) neurotoxicity to striatal dopamine neurons in mice

1. MPTP given to mice in 4 daily doses (20 mg/kg s.c.) resulted in 56–70% depletion of striatal dopamine 1 week after the last dose.

2. Pretreatment with deprenyl or MD 240928, selective inhibitors of monoamine oxidase type B, or with amfonelic acid or nomifensine, selective inhibitors of dopamine uptake, prevented the depletion of striatal dopamine. In contrast, pretreatment with -methyltyrosine, Ro 4-1284 or haloperidol did not prevent the depletion of striatal dopamine by MPTP.

3. The results are compatible with the view that dopamine itself is not involved in the neurotoxic effect of MPTP but that MPP+, a metabolite of MPTP formed by MAO-B and accumulated by the dopamine uptake carrier, is responsible for the neurotoxicity.     

Upregulation of striatal D2 receptors in the MPTP-treated vervet monkey is reversed by grafts of fetal ventral mesencephalon: an autoradiographic study

Although neural transplantation holds promise as a treatment for Parkinson's disease, parkinsonian primates have generally exhibited inconsistent and incomplete recovery of motor functions following intrastriatal grafting of fetal ventral mesencephalon. One possible contributing factor to this variable response is lack of appropriate integration of donor neurons with host striatal circuitry with the result that there is insufficient dopamine release and postsynaptic dopamine receptor activation. This issue was examined by measuring the effect of transplanting fetal ventral mesencephalon to the striatum of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated (MPTP) monkeys on striatal D2 receptor binding. One year after receiving MPTP, D2 receptor binding was upregulated in the dorsal and ventral striatum of African green monkeys. Grafting of fetal ventral mesencephalon to the dorsal striatum of MPTP-treated monkeys 9 months before sacrifice, eliminated the D2 receptor upregulation in dorsal, but not ventral, region. Dopamine concentration in dorsal striatum of grafted MPTP-treated monkeys was significantly higher than in that region of MPTP-treated non-grafted monkeys. In addition, dopamine concentration was significantly higher in dorsal compared to ventral striatum of grafted MPTP-treated monkeys. These data, in addition to those from a previous autoradiographic study on dopamine uptake site density in these monkeys, strongly supports the hypothesis that ectopically placed ventral mesencephalon not only produces, but maintains the release of sufficient levels of dopamine to restore postsynaptic dopamine transmission in regions influenced by graft-derived dopamine.     

In vitro effects of catecholamines and catecholestrogens on brain tyrosine hydroxylase activity and kinetics in the female catfish Heteropneustes fossilis

Effects of catecholamines and catecholestrogens on tyrosine hydroxylase (TH) activity and kinetics were investigated in the telencephalon and hypothalamus of female Heteropneustes fossilis in gonad quiescent (resting) and recrudescent (preparatory) phases. Dopamine, noradrenaline and adrenaline and the catecholestrogen, 2-hydroxyestradiol-17 beta inhibited TH activity in a concentration-dependent manner in both resting and preparatory phases, with a higher effect in the resting phase. Two- methoxyestradiol-17 beta did not alter TH activity in any season. The catecholamines inhibited TH in a competitive manner increasing apparent K(m) values significantly without altering the apparent V(max). Two-hydroxyestradiol-17 beta inhibited significantly the enzyme in a noncompetitive manner and decreased apparent V(max) without altering apparent K(m) values. The apparent K(i) is higher for dopamine than noradrenaline or adrenaline. The apparent K(i) for 2-hydroxyestradiol-17 beta is not significantly different from that of noradrenaline. The present results suggest an interaction between oestradiol-17beta (E2) and catecholamine metabolism at the level of tyrosine hydroxylation and E2 effects on catecholamines may be mediated through its 2-hydroxylation.     

Quantitative analyses of GFRalpha-1 and GFRalpha-2 mRNAs and tyrosine hydroxylase protein in the nigrostriatal system reveal bilateral compensatory changes following unilateral 6-OHDA lesions in the rat

Copy numbers of mRNAs for GFRalpha-1 and GFRalpha-2, the preferred receptors for glial cell line-derived neurotrophic factor (GDNF) and neurturin (NTN) were determined by real-time quantitative RT-PCR (QRT-PCR). Receptor expression was assessed in striatum (ST) and substantia nigra (SN) of normal rats and rats acutely or progressively lesioned by 6-OHDA injected into the medial forebrain bundle or ST, respectively. GFRalpha-1 mRNA was clearly detected in normal ST. In normal SN, significantly higher expression of both receptors was observed. At 4 weeks after acute lesion, GFRalpha-2 mRNA was markedly decreased in SN bilaterally, whereas GFRalpha-1 mRNA in SN and ST was not affected. A progressive lesion resulted in a progressive decrease of GFRalpha1 mRNA in ST bilaterally. In SN, levels of GFRalpha-1 mRNA were not significantly affected by a progressive lesion, whereas GFRalpha-2 mRNA was markedly decreased bilaterally. Quantitative western blotting standardized against tyrosine hydroxylase (TH) protein from PC12 cells revealed the expected decrease in TH protein in lesioned SN, but also significant increases in TH protein in contralateral, unlesioned SNs at 4 weeks after both acute and progressive lesions. These data suggest that previously unrecognized compensatory changes in the nigrostriatal system occur in response to unilateral dopamine depletion. Since the changes observed in receptor expression did not always parallel loss of dopamine neurons, cells in addition to the nigral dopamine neurons appear to be affected by a 6-OHDA insult and are potential targets for the neurotrophic factors, GDNF and NTN.     

Neurotensin terminals form synapses primarily with neurons lacking detectable tyrosine hydroxylase immunoreactivity in the rat substantia nigra and ventral tegmental area.

A light and electron microscopic double antigen localization technique was employed to examine the fine structural relationship between neurotensin-containing axon terminals and dopaminergic neurons in the substantia nigra and ventral tegmental area of the rat. At the light microscopic level, neurotensin-immunoreactive terminals were densely distributed throughout the substantia nigra pars compacta and ventral tegmental area in close proximity to tyrosine hydroxylase-immunoreactive somata and dendrites. On electron microscopic examination, direct synaptic connections were identified between neurotensin-immunoreactive axon terminals and tyrosine hydroxylase-immunopositive perikarya and dendrites. However, only 8.2% and 8.8% of the neurotensin-immunoreactive axonal profiles detected in the substantia nigra and ventral tegmental area, respectively, were found in direct apposition with tyrosine hydroxylase-immunostained elements. In turn, only 9.3% and 10.0% of tyrosine hydroxylase immunoreactive dendrites sampled from the substantia nigra and ventral tegmental area, respectively, were seen in contact with neurotensin immunopositive axon terminals. However, neurotensin-immunoreactive and tyrosine hydroxylase-immunolabelled elements were frequently identified in close anatomical proximity (less than 5 microns) to one another. These results are interpreted in light of the selective association of neurotensin receptors with dopaminergic neurons in the substantia nigra and ventral tegmental area to suggest a predominantly parasynaptic mechanism of action for neurotensin in the ventral midbrain.     

Differential activation of PKC delta in the substantia nigra of rats following striatal or nigral 6-hydroxydopamine lesions

Parkinsonian neurodegeneration is associated with heightened levels of oxidative stress and the activation of apoptotic pathways. In an in vitro cellular model, we reported that 6-hydroxydopamine (6-OHDA) induces apoptotic cell death via the induction of mitochondrial dysfunction, the activation of caspase 3 and the consequent proteolytic activation of the redox-sensitive kinase, protein kinase C (PKC)δ, in PC12 cells. Here we have investigated the involvement of PKCδ in 6-OHDA-induced cell death in vivo . The nigrostriatal pathway of rats was lesioned by unilateral infusion of 6-OHDA into either the striatum or substantia nigra pars compacta (SNpc). Infusion into the SNpc resulted in rapid loss of tyrosine hydroxylase (TH)-positive cells (87% decrease after 4 days), consistent with a necrotic-like mode of cell death. In contrast, striatal infusion initiated a slower, progressive decline in TH immunoreactivity (25% decrease in the SNpc after 4 days); cell appearance was characteristic of apoptosis. This is consistent with a transient increase in active caspase 3 immunoreactivity at 4 days post-infusion, and a concomitant proteolytic activation of PKCδ in the SNpc of striatal-lesioned rats. Cleavage of PKCδ did not occur in the striatum or cerebellum of lesioned animals, or in the SNpc of sham-operated controls. No increase in caspase 3 immunoreactivity or proteolytic activation of PKCδ was detected in nigral-lesioned rats. These results suggest that after 6-OHDA infusion into the striatum, retrograde neurotoxicity induces caspase 3-dependent PKCδ proteolytic activation in the cell bodies of the SNpc, implicating this kinase in the neurodegenerative process.     

Platelet monoamine oxidase activity in relation to alleles of dopamine D4 receptor and tyrosine hydroxylase genes

Human personality characteristics and vulnerability to psychopathology are to a large extent dependent upon genetic factors which have yet to be fully defined. The allele distribution of the dopamine D4 receptor (D4DR) and thrombocyte monoamine oxidase (trbc MAO) activity have both been associated with personality traits which are supposedly related, namely 'sensation seeking’according to Zuckerman and‘novelty seeking’according to Cloninger, respectively. In this report, the D4DR allele distribution and trbc MAO activity were studied in 31 psychiatric patients and 21 control subjects. Trbc MAO activity is a biochemical marker of personality that has been shown to be under strong genetic influence. However, no association between the D4DR alleles and trbc MAO could be observed in this material. To our knowledge, this is the first report comparing these two markers, and based upon the results obtained, we speculate that they may be connected with different types of overlapping personality characteristics. The allele distribution of the tyrosine hydroxylase (TH) gene was also determined. TH is the rate-limiting enzyme in the biosynthesis of catecholamines, and it is believed to be involved in different kinds of psychopathology. No covariation between TH gene alleles and trbc MAO activity or D4DR alleles was observed in this material.