Tyrosine Hydroxylase Immunohistochemistry Research Articles

Fos expression in pontomedullary catecholaminergic cells following rapid eye movement sleep-like episodes elicited by pontine carbachol in urethane-anesthetized rats

Pontine noradrenergic neurons of the locus coeruleus (LC) and sub-coeruleus (SubC) region cease firing during rapid eye movement sleep (REMS). This plays a permissive role in the generation of REMS and may contribute to state-dependent modulation of transmission in the CNS. Whether all pontomedullary catecholaminergic neurons, including those in the A1/C1, A2/C2 and A7 groups, have REMS-related suppression of activity has not been tested. We used Fos protein expression as an indirect marker of the level of neuronal activity and linear regression analysis to determine whether pontomedullary cells identified by tyrosine hydroxylase (TH) immunohistochemistry have reduced Fos expression following REMS-like state induced by pontine microinjections of a cholinergic agonist, carbachol in urethane-anesthetized rats. The percentage of Fos-positive TH cells was negatively correlated with the cumulative duration of REMS-like episodes induced during 140 min prior to brain harvesting in the A7 and rostral A5 groups bilaterally ( P<0.01 for both), and in SubC neurons on the side opposite to carbachol injection ( P<0.05). Dorsal medullary A2/C2 neurons did not exhibit such correlation, but their Fos expression (and that in A7, rostral A5 and SubC neurons) was positively correlated with the duration of the interval between the last REMS-like episode and the time of perfusion ( P<0.05). In contrast, neither of these correlations was significant for A1/C1 or caudal A5 neurons. These findings suggest that, similar to the prototypic LC neurons, neurons of the A7, rostral A5 and A2/C2 groups have reduced or abolished activity during REMS, whereas A1/C1 and caudal A5 neurons do not have this feature. The reduced activity of A2/C2, A5 and A7 neurons during REMS, and the associated decrements in norepinephrine release, may cause state-dependent modulation of transmission in brain somato- and viscerosensory, somatomotor, and cardiorespiratory pathways.

Generation of a α-synuclein-based rat model of Parkinson's disease

Two missense mutations (A30P and A53T) in the gene for alpha-synuclein (α-syn) cause familial Parkinson's disease (PD) in a small cohort. There is increasing evidence to propose that abnormal metabolism and accumulation of α-syn in dopaminergic neurons play a role in the development of familial as well as sporadic PD. The complexity of the mechanisms underlying α-syn-induced neurotoxicity, however, has made difficult the development of animal models that faithfully reproduce human PD pathology. We now describe and characterize such a model, which is based on the stereotaxic injection into rat right substantia nigra pars compacta of the A30P mutated form of α-syn fused to a protein transduction domain (TAT). The TAT sequence allows diffusion of the fusion protein across the neuronal plasma membrane and results in a localized dopaminergic loss. Dopaminergic cell loss was evaluated both by tyrosine hydroxylase immunohistochemistry and by HPLC analysis of dopamine and its catabolite 3,4 dihydroxyphenylacetic acid. Infusion of TAT-α-synA30P induced a significant 26% loss in dopaminergic neurons. This dopaminergic loss was accompanied by a time-dependent impairment in motor function, evaluated utilizing the rotarod and footprint tests. In comparison to chemical neurotoxin-based (e.g. 6-hyroxydopamine, MPTP) animal models of PD, the α-syn-based PD animal model offers the advantage of mimicking the early stages and slow development of the human disease and should prove valuable in assessing specific aspects of PD pathogenesis in vivo and in developing new therapeutic strategies.

The Hypothalamic Supraoptic and Paraventricular Nuclei of the Echidna and Platypus

The monotremes are an intriguing group of mammals that have major differences in their reproductive physiology and lactation from therian mammals. Monotreme young hatch from leathery skinned eggs and are nourished by milk secreted onto areolae rather than through nipples. Parturition and lactation are in part controlled through the paraventricular and supraoptic nuclei of the hypothalamus. We have used Nissl staining, enzyme histochemistry, immunohistochemistry for tyrosine hydroxylase, calbindin, oxytocin, neurophysin and non-phosphorylated neurofilament protein, and carbocyanine dye tracing techniques to examine the supraoptic and paraventricular nuclei and the course of the hypothalamo-neurohypophysial tract in two monotremes: the short-beaked echidna (Tachyglossus aculeatus) and the platypus (Ornithorhynchus anatinus). In both monotremes, the supraoptic nucleus consisted of loosely packed neurons, mainly in the retrochiasmatic position. In the echidna, the paraventricular nucleus was quite small, but had similar chemoarchitectural features to therians. In the platypus, the paraventricular nucleus was larger and appeared to be part of a stream of magnocellular neurons extending from the paraventricular nucleus to the retrochiasmatic supraoptic nucleus. Immunohistochemistry for non-phosphorylated neurofilament protein and carbocyanine dye tracing suggested that hypothalamo-neurohypophysial tract neurons in the echidna lie mainly in the retrochiasmatic supraoptic and lateral hypothalamic regions, but most neurophysin and oxytocin immunoreactive neurons in the echidna were found in the paraventricular, lateral hypothalamus and supraoptic nuclei and most oxytocinergic neurons in the platypus were distributed in a band from the paraventricular nucleus to the retrochiasmatic supraoptic nucleus. The small size of the supraoptic nucleus in the two monotremes might reflect functional aspects of monotreme lactation.

PGE2 receptor EP1 renders dopaminergic neurons selectively vulnerable to low‐level oxidative stress and direct PGE2 neurotoxicity

Oxidative stress and increased cyclooxygenase-2 (COX-2) activity are both implicated in the loss of dopaminergic neurons from the substantia nigra (SN) in idiopathic Parkinson's disease (PD). Prostaglandin E(2) (PGE(2)) is one of the key products of COX-2 activity and PGE(2) production is increased in PD. However, little is known about its role in the selective death of dopaminergic neurons. Previously, we showed that oxidative stress evoked by low concentrations of 6-hydroxydopamine (6-OHDA) was selective for dopaminergic neurons in culture and fully dependent on COX-2 activity. We postulated that this loss was mediated by PGE(2) acting through its receptors, EP1, EP2, EP3, and EP4. Using double-label immunohistochemistry for specific EP receptors and tyrosine hydroxylase (TH), we identified EP1 and EP2 receptors on dopaminergic neurons in rat SN. EP2 receptors were also found in non-dopaminergic neurons of this nucleus, as were EP3 receptors, whereas the EP4 receptor was absent. PGE(2), 16-phenyl tetranor PGE(2) (a stable synthetic analogue), and 17-phenyl trinor PGE(2) (an EP1 receptor-selective agonist) were significantly toxic to dopaminergic cells at nanomolar concentrations; EP2- and EP3-selective agonists were not. We challenged dopaminergic neurons in embryonic rat mesencephalic primary neuronal cultures and tested whether these receptors mediate selective 6-OHDA toxicity. The nonselective EP1-3 receptor antagonist AH-6809 and two selective EP1 antagonists, SC-19220 and SC-51089, completely prevented the 40%-50% loss of dopaminergic neurons caused by exposure to 5 muM 6-OHDA. Together, these results strongly implicate PGE(2) activation of EP1 receptors as a mediator of selective toxicity in this model of dopaminergic cell loss.

Distribution and morphology of putative catecholaminergic and serotonergic neurons in the brain of the greater canerat, Thryonomys swinderianus

The distribution, morphology and nuclear subdivisions of the putative catecholaminergic and serotonergic systems within the brain of the greater canerat (sometimes spelt cane rat) were identified following immunohistochemistry for tyrosine hydroxylase and serotonin. The aim of the present study was to investigate possible differences in the complement of nuclear subdivisions of these systems when comparing those of the greater canerat with reports of these systems in other rodents. The greater canerat was chosen for investigation as it is a large rodent (around 2.7 kg body mass) and has an average brain mass of 13.75 g, more than five times larger than that of the laboratory rat. The greater canerats used in the present study were caught from the wild, which is again another contrast to the laboratory rat. While these differences, especially that of size, may lead to the prediction of significant differences in the nuclear complement of these systems, we found that all nuclei identified in both systems in the laboratory rat and other rodents in several earlier studies had direct homologs in the brain of the greater canerat. Moreover, there were no additional nuclei in the brain of the greater canerat that are not found in the laboratory rat or other rodents. It is noted that the locus coeruleus of the laboratory rat differs in appearance to that reported for several other rodent species. The greater canerat is phylogenetically distant from the laboratory rat, but still a member of the order Rodentia. Thus, changes in the nuclear organization of these systems appears to demonstrate a form of constraint related to the phylogenetic level of the order.

Development of neurotransmitter specificity in sympathetic ganglionic neurons

The neurotransmitter features in sympathetic neurons are subject to change during development (Masliukov, Timmermans, 2004). For better understanding the neuroplasticity of sympathetic neurons during early postnatal ontogenesis, the purpose of this study was to investigate the development of the catecholaminergic, cholinergic and peptidergic phenotypes in the superior cervical ganglion (SCG) and celiac ganglia (CG) neurons. The present study was performed in rats of different ages (newborn, 10-, 20-, 30-, 60-day-old) under a lethal dose of sodium pentobarbital (Nembutal, 300 mg/kg, i.p.). Immunohistochemistry for tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), vasoactive intestinal (poly)peptide (VIP), neuropeptide Y (NPY), somatostatin (SOM), substance P (SP) and histochemical method for NADPH-diaphorase was applied. The results obtained indicate that the majority of the sympathetic neurons in the SCG and CG were TH-positive from birth onwards. A large part of sympathetic neurons also contained NPY. The percentage of neurons containing TH and NPY invariably increased with age up to 60 days postnatally. The percentage of NPY neurons was lower in the CG in accordance with SCG. Only few neurons contained VIP- and ChAT in the sympathetic ganglia in all ages under study. The population of SOM-positive neurons was substantially greater in the CG in comparison with the SCG. The percentage of SOM-positive cells in the SCG significantly decreased from birth onwards and we observed only single cells in 10-day-old and older animals. Meanwhile, the proportion of SOM-immunoreactive cells in the CG was not change during the development. SP was found in single neurons in the CG and SCG. NADPH-diaphorase positive cells were absent in the SCG and CG. In conclusion, the neurotransmitter composition in the SCG, CG and IG is present even in newborn animals. However, these ganglia develop heterochronously. Finally, the neurotransmitter set becomes complete by the second month of life.

High frequency stimulation of the subthalamic nucleus improves speed of locomotion but impairs forelimb movement in Parkinsonian rats

The subthalamic nucleus (STN) plays an important role in motor and non-motor behavior in Parkinson’s disease, but its involvement in gait functions is largely unknown. In this study, we investigated the role of the STN on gait in a rat model of PD using the CatWalk method. Parkinsonian rats received bilateral high frequency stimulation (HFS) with different stimulation amplitudes of the STN. Rats were rendered parkinsonian by bilateral injections of 6-hydroxydopamine (6-OHDA) into the striatum. One group of 6-OHDA animals was implanted bilaterally with stimulation electrodes at the level of the STN. Stimulations were performed at 130 Hz (frequency), 60 μs (pulse width) and varying amplitudes of 0, 3, 30 and 150 μA. Rats were evaluated in an automated quantitative gait analysis method (CatWalk method). After behavioral evaluations, rats were killed and the brains processed for histological stainings to determine the impact of the dopaminergic lesion (tyrosine hydroxylase immunohistochemistry) and the localization of the electrode tip (hematoxylin–eosin histochemistry). Results show that bilateral 6-OHDA infusion significantly decreased (70%) the number of dopaminergic cells in the substantia nigra pars compacta (SNc). Due to 6-OHDA treatment, the gait parameters changed considerably. There was a general slowness. The most pronounced effects were seen at the level of the hind paws. Due to implantation of STN electrodes the step pattern changed. STN electrical stimulation improved the general slowness but induced slowing of the forelimb movement. Furthermore, we found that HFS with a medium amplitude significantly changed speed, the so-called dynamic aspect of gait. The static features of gait were only significantly influenced with low amplitude. Remarkably, STN stimulation affected predominantly the forepaws/limbs.

L-Dopa treatment abolishes the numerical increase in striatal dopaminergic neurons in parkinsonian monkeys

The striatum harbors a population of dopaminergic interneurons that increases in number in animal models of Parkinson's disease (PD), presumably to compensate for dopamine (DA) depletion. The purpose of the present study was to determine the fate of striatal dopaminergic neurons in parkinsonian monkeys in which striatal DA depletion had been alleviated by systemic administration of l-dopa. The number of striatal dopaminergic neurons, visualized with tyrosine hydroxylase (TH) immunohistochemistry, was measured in three groups of cynomolgus ( Macaca fascicularis) monkeys: (1) normal untreated monkeys; (2) monkeys rendered parkinsonian following systemic injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), but otherwise untreated; and (3) MPTP-intoxicated monkeys that received oral l-dopa on a chronic basis. In agreement with previous studies, the number of striatal TH-positive (TH+) neurons in l-dopa-free parkinsonian monkeys was significantly higher ( p < 0.05) than in normal (non-parkinsonian) monkeys. However, this increase was abolished in parkinsonian monkeys that received l-dopa treatment. In fact, the number of striatal TH+ neurons in l-dopa-treated parkinsonian monkeys was not significantly different ( p > 0.05) from values obtained in normal monkeys. These findings suggest that the DA concentration regulates the numerical density of this ectopic neuronal population, a phenomenon that is more likely the result of a shift in the phenotype of preexistent striatal interneurons rather than the recruitment of newborn neurons that would eventually develop a DA phenotype. Our data also reinforce the hypothesis that striatal TH+ neurons act as local DA source and, as such, are part of a compensatory mechanism that could be artificially enhanced to alleviate or delay PD symptoms.

Distribution and morphology of catecholaminergic and serotonergic neurons in the brain of the highveld gerbil, Tatera brantsii

The distribution, morphology and nuclear subdivisions of the putative catecholaminergic and serotonergic systems within the brain of the highveld gerbil were identified following immunohistochemistry for tyrosine hydroxylase and serotonin. The aim of the present study was to investigate possible differences in the complement of nuclear subdivisions of these systems when comparing those of the highveld gerbil with those of the laboratory rat. The highveld gerbil was chosen as it is relatively closely related to the laboratory rat, but the Gerbillinae and Murinae lineages diverged over 20 million years ago. Moreover, even though brain sizes are similar, the life history and phenotypes between these two species are substantially different. The gerbils used in the present study were caught from the wild, which is again another contrast to the laboratory rat. While these differences may lead to the prediction of significant differences in the nuclear complement of these systems, we found that all nuclei identified in both systems in the laboratory rat in several earlier studies had direct homologs in the brain of the highveld gerbil. Moreover, there were no additional nuclei in the brain of the highveld gerbil that are not found in the laboratory rat. The only discernable difference between the two species was a greater density and number of catecholaminergic neurons in the olfactory bulb of the highveld gerbil. Thus, the evolution of nuclear parcellation in these systems appears to demonstrate a form of phylogenetic constraint related to the order Rodentia.

Positional relationship between natural killer cells and distribution of sympathetic nerves in decidualized mouse uterus.

Uterine natural killer (uNK) cells are the most abundant leukocytes in pre-implantation endometrium and early pregnancy deciduas in humans and rodents. They are associated with structural changes in maternal spiral arteries but regulation of their recruitment and activation is incompletely understood. The major subpopulation of uNK cells in humans expresses CD56, the neural cell adhesion molecule (NCAM)-1 while their counterpart in mouse expresses asialoGM1, a brain ganglioside. Sympathetic nerves express NCAM-1 which mediates homotypic binding. Sympathetic fibers innervate the mesometrial vasculature but their relationship to the myometrial and decidual uNK cell recruitment is unknown. The present study aims to explore positional relationship between natural killer cells and distribution of nerves in decidualized mouse uterus. Immunohistochemistry and mRNA expression for the enzyme tyrosine hydroxylase were used to map sympathetic nerve fibre distribution within C57BL/6 implantation sites and to address a relationship with uNK cells. Tyrosine hydroxylase positive neurons were identified in the mesometrium closely associated with uterine arteries. Staining became gradually vanished as the nerves crossed the myometrium and entered the decidualized uterus. No neuronal stain was associated with the spiral arteries. Periodic Acid Schiff's reactive uNK cells were absent from the mesentery, but abundant in decidua basalis where they are associated with non-innervated vessels. Data suggest that the recruitment of uNK progenitor cells to the uterus is unlikely to be dependent on signaling by the sympathetic nervous system.

Histological Validation of Dopamine Transporter PET Images in the Rat 6-OHDA Lesion Model of Parkinson's Disease

Positron emission tomography (PET) imaging of dopamine reuptake transporters (DAT) in the rat 6-hydoxydopamine (6-OHDA) lesion model of Parkinson's disease (PD) is a clinically translatable biomarker that could be used in the discovery of novel, disease-modifying PD therapeutics.Careful histological validation of DAT PET images are critical steps in overcoming clinical imaging artifacts seen in PD patients undergoing dopamine replacement therapy. In this report, we describe the use of quantitative DAT autoradiography and tyrosine hydroxylase immunohistochemistry methods using adjacent cryosections to provide histological validation of DAT PET images and verify the loss of DAT binding sites with reduced survival of dopaminergic neurons in the nigrostriatal pathway. (The J Histotechnol 30:123, 2007)Submitted March 1, 2007; accepted with revisions April 3, 2007

Distribution and morphology of putative catecholaminergic and serotonergic neurons in the medulla oblongata of a sub-adult giraffe, Giraffa camelopardalis

The current study details the nuclear parcellation and appearance of putative catecholaminergic and serotonergic neurons within the medulla oblongata of a sub-adult giraffe, using immunohistochemistry for tyrosine hydroxylase and serotonin. We hypothesized that the unusual phenotype of the giraffe, this being the long neck and potential axonal lengthening of these neurons, may pose specific problems in terms of the efficient functioning of these systems, as several groups of catecholaminergic and serotonergic neurons, especially of the medulla, are known to project to the entire spinal cord. This specific challenge may lead to observable differences in the nuclear parcellation and morphology of these systems in the giraffe. Our personal observations in the giraffe reveal that, as with other Artiodactyls, the spinal cord extends to the caudal end of the sacral vertebrae. Within the giraffe medulla we found evidence for five putative catecholaminergic (neurons containing tyrosine hydroxylase) and five serotonergic nuclei. In terms of both morphological appearance of the neurons and nuclear parcellation we did not find any evidence for features that may be considered affected by the phenotype of the giraffe. The nuclear parcellation and appearance of both the putative catecholaminergic and serotonergic systems in the medulla of the giraffe studied are strikingly similar to that seen in previous studies of other Artiodactyls. We interpret these findings in terms of a growing literature detailing order specific phylogenetic constraints in the evolution of these neuromodulatory systems.

Developmental changes of the adrenergic network in the myenteric plexus of the porcine small bowel

Knowledge about the foetal development of the normal enteric nervous system (ENS) is crucial for the understanding of congenital and acquired functional abnormalities of the gut. The ENS is the largest and most complex division of the peripheral nervous system and consists of intrinsic and extrinsic components. Although previous studies have described sympathetic innervation of the myenteric plexus, little is known regarding its age-related changes. The aim of this study was to investigate the age-related changes in the sympathetic innervation of the myenteric plexus. Whole mount and paraffin sections of the small bowel specimens from six different age groups (60 and 90 days gestation; newborn; 4 and 12 weeks old; and adult) were stained using tyrosine hydroxylase immunohistochemistry. Specimens were then analysed using fluorescence and laser scanning microscopy in detail. The tyrosine hydroxylase positive nerve fibres were first seen within the myenteric plexus at 90 days of gestation (E90). There was a significant increase in nerve fibres and varicosities observed from E90 to 12 weeks of age and stabilisation thereafter. The degree of varicosities around the ganglia, clearly seen on the whole-mount preparations, was also noted to increase up to 12 weeks of age, after which time there was no general variation noted into adulthood. Our findings show, for the first time, that sympathetic innervation of the myenteric plexus starts in the last quarter of gestation and continues till 12 weeks of age. Segmental sympathetic denervation, following bowel resection and anastomosis, during this developmental period may explain the motility dysfunction seen in newborn infants operated for necrotising enterocolitis, bowel atresia and Hirschsprung's disease.

Peripheral iron dextran induced degeneration of dopaminergic neurons in rat substantia nigra

Iron accumulation is considered to be involved in the pathogenesis of Parkinson's disease. To demonstrate the relationship between peripheral iron overload and dopaminergic neuron loss in rat substantia nigra (SN), in the present study we used fast cyclic voltammetry, tyrosine hydroxylase (TH) immunohistochemistry, Perls’ iron staining, and high performance liquid chromatography-electrochemical detection to study the degeneration of dopaminergic neurons and increased iron content in the SN of iron dextran overloaded animals. The findings showed that peripheral iron dextran overload increased the iron staining positive cells and reduced the number of TH-immunoreactive neurons in the SN. As a result, dopamine release and content, as well as its metabolites contents were decreased in caudate putamen. Even more dramatic changes were found in chronic overload group. These results suggest that peripheral iron dextran can increase the iron level in the SN, where excessive iron causes the degeneration of dopaminergic neurons. The chronic iron overload may be more destructive to dopaminergic neurons than the acute iron overload.

Changes in brain functional activation during resting and locomotor states after unilateral nigrostriatal damage in rats

To evaluate functional neuronal compensation after partial damage to the nigrostriatal system, we lesioned rats unilaterally in the striatum with 6-hydroxydopamine. Five weeks later, cerebral perfusion was mapped at rest or during treadmill walking using [14C]-iodoantipyrine. Regional CBF-related tissue radioactivity (CBF-TR) was quantified by autoradiography and analyzed by statistical parametric mapping and region-of- interest analysis. Lesions were confirmed by tyrosine hydroxylase immunohistochemistry and changes in rotational locomotor activity.Functional compensations were bilateral and differed at rest and during treadmill walking. Consistent with the classic view of striatopallidal connections, CBF-TR of lesioned compared to sham-lesioned rats increased in the ipsilateral subthalamic nucleus (STN) and internal globus pallidus, and decreased in the striatum and external globus pallidus. Contrary to the classic view, CBF-TR increased in the ipsilateral ventral lateral, ventral anterior thalamus and motor cortex, as well as in the central medial thalamus, midline cerebellum, and contralateral STN.During walking, perfusion decreased in lesioned compared to sham-lesioned rats across the ipsilateral striato-pallidal-thalamic-cortical motor circuit. Compensatory increases were seen bilaterally in the ventromedial thalamus and red nucleus, in the contralateral STN, anterior substantia nigra, subiculum, motor cortex, and in midline cerebellum. Enhanced recruitment of associative sensory areas was noted cortically and subcortically.Future models of compensatory changes after nigrostriatal damage need to address the effects of increased neural activity by residual dopaminergic neurons, interhemispheric interactions and differences between resting and locomotor states. Identification of sites at which functional compensation occurs may define useful future targets for neurorehabilitative or neurorestorative interventions in Parkinson's disease.

Sex differences in catechol contents in the olfactory bulb of control and unilaterally deprived rats

The dopaminergic system plays important roles in the modulation of olfactory transmission. The present study examines the distribution of dopaminergic cells and the content of dopamine (DA) and its metabolites in control and deprived olfactory bulbs (OB), focusing on the differences between sexes. The content of DA and of its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), were measured by HPLC. The morphology and distribution of dopaminergic neurons were studied using tyrosine hydroxylase (TH) immunohistochemistry. Cells were typified with TH-parvalbumin, TH-cholecystokinin or TH-neurocalcin double-immunofluorescence assays. Biochemical analyses revealed sex differences in the content of DA and of its metabolites. In normal conditions, the OBs of male rats had higher concentrations of DA, DOPAC and HVA than the OBs of females. The immunohistochemical data pointed to sex differences in the number of TH-immunopositive cells (higher in male than in female rats). Colocalization analyses revealed that dopaminergic cells constitute a different cell subpopulation from those labelled after parvalbumin, cholecystokinin or neurocalcin immunostaining. Unilateral olfactory deprivation caused dramatic alterations in the dopaminergic system. The DA content and the density of dopaminergic cells decreased, the contents of DA and DOPAC as well as TH immunoreactivity were similar in deprived males and females and, finally, the metabolite/neurotransmitter ratio increased. Our results show that the dopaminergic modulation of olfactory transmission seems to differ between males and females and that it is regulated by peripheral olfactory activity. A possible role of the dopaminergic system in the sexually different olfactory sensitivity, discrimination and memory is discussed.

Structure and chemical organization of the accessory olfactory bulb in the goat

The structure and chemical composition of the accessory olfactory bulb (AOB) were examined in male and female goats. Sections were subjected to either Nissl staining, Klüver-Barrera staining, lectin histochemistry, or immunohistochemistry for nitric oxide synthase (NOS), neuropeptide Y (NPY), tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), and glutamic acid decarboxylase (GAD). The goat AOB was divided into four layers: the vomeronasal nerve layer (VNL), glomerular layer (GL), mitral/tufted (M/T) cell layer (MTL), and granule cell layer (GRL). Quantitative and morphometric analyses indicated that a single AOB contained 5,000-8,000 putative M/T cells with no sex differences, whereas the AOB was slightly larger in males. Of the 21 lectins examined, 7 specifically bound to the VNL and GL, and 1 bound not only to the VNL, but also to the MTL and GRL. In either of these cases, no heterogeneity of lectin staining was observed in the rostrocaudal direction. NOS-, TH-, DBH-, and GAD-immunoreactivity (ir) were observed in the MTL and GRL, whereas NPY-ir was present only in the GRL. In the GL, periglomerular cells with GAD-ir were found in abundance, and a subset of periglomerular cells containing TH-ir was also found. Double-labeling immunohistochemistry revealed that virtually all periglomerular cells containing TH-ir were colocalized with GAD-ir.

Sept4, a Component of Presynaptic Scaffold and Lewy Bodies, Is Required for the Suppression of α-Synuclein Neurotoxicity

In Parkinson disease (PD), alpha-synuclein aggregates called Lewy bodies often involve and sequester Septin4 (Sept4), a polymerizing scaffold protein. However, the pathophysiological significance of this phenomenon is unclear. Here, we show the physiological association of Sept4 with alpha-synuclein, the dopamine transporter, and other presynaptic proteins in dopaminergic neurons; mice lacking Sept4 exhibit diminished dopaminergic neurotransmission due to scarcity of these presynaptic proteins. These data demonstrate an important role for septin scaffolds in the brain. In transgenic mice that express human alpha-synuclein(A53T) (a mutant protein responsible for familial PD), loss of Sept4 significantly enhances neuropathology and locomotor deterioration. In this PD model, insoluble deposits of Ser129-phosphorylated alpha-synuclein(A53T) are negatively correlated with the dosage of Sept4. In vitro, direct association with Sept4 protects alpha-synuclein against self-aggregation and Ser129 phosphorylation. Taken together, these data show that Sept4 may be involved in PD as a dual susceptibility factor, as its insufficiency can diminish dopaminergic neurotransmission and enhance alpha-synuclein neurotoxicity.

Intrastriatal gene transfer of vascular endothelial growth factor rescues dopaminergic neurons in a rat Parkinson’s disease model

To examine the ability of intrastriatal gene transfer of vascular endothelial growth factor 165 mediated by adenoviral vector to rescue dopaminergic neurons in a rat model of Parkinson's disease (PD), we constructed recombinant replication-deficient adenoviral vectors carrying the gene of VEGF165 (Ad-VEGF), and injected Ad-VEGF (or Ad-LacZ and PBS as controls) into the striatum of rats 7 days after the lesion by 6-hydroxydopamine. The rat rotational behavior analysis and tyrosine hydroxylase (TH) immunohistochemistry were performed to assess the change of dopaminergic neurons. Our results showed that the rats receiving Ad-VEGF injection displayed a significant improvement in apomorphine-induced rotational behavior and a significant preservation of TH-positive neurons and fibers compared with control animals. It is concluded that intrastriatal gene transfer by Ad-VEGF may rescue the dopaminergic neurons from degeneration in a rat model of PD.

Chicken lateral septal organ and other circumventricular organs form in a striatal subdomain abutting the molecular striatopallidal border

The avian lateral septal organ (LSO) is a telencephalic circumventricular specialization with liquor-contacting neurons (Kuenzel and van Tienhoven [1982] J. Comp. Neurol. 206:293-313). We studied the topological position of the chicken LSO relative to molecular borders defined previously within the telencephalic subpallium (Puelles et al. [2000] J. Comp. Neurol. 424:409-438). Differential expression of Dlx5 and Nkx2.1 homeobox genes, or the Shh gene encoding a secreted morphogen, allows distinction of striatal, pallidal, and preoptic subpallial sectors. The chicken LSO complex was characterized chemoarchitectonically from embryonic to posthatching stages, by using immunohistochemistry for calbindin, tyrosine hydroxylase, NKX2.1, and BEN proteins and in situ hybridization for Nkx2.1, Nkx2.2, Nkx6.1, Shh, and Dlx5 mRNA. Medial and lateral parts of LSO appear, respectively, at the striatal part of the septum and adjacent bottom of the lateral ventricle (accumbens), in lateral continuity with another circumventricular organ that forms along a thin subregion of the entire striatum, abutting the molecular striatopallidal boundary; we called this the "striatopallidal organ" (SPO). The SPO displays associated distal periventricular cells, which are lacking in the LSO. Moreover, the SPO is continuous caudomedially with a thin, linear ependymal specialization found around the extended amygdala and preoptic areas. This differs from SPO and LSO in some molecular aspects. We tentatively identified this structure as being composed of an "extended amygdala organ" (EAO) and a "preoptohypothalamic organ" (PHO). The position of LSO, SPO, EAO, and PHO within a linear Dlx5-expressing ventricular domain that surrounds the Nkx2.1-expressing pallidopreoptic domain provides an unexpected insight into possible common and differential causal mechanisms underlying their formation.