Catecholaminergic Neurons Research Articles

Mapping of the excitatory, inhibitory, and modulatory afferent projections to the anatomically defined active expiratory oscillator in adult male rats.

The lateral parafacial region (pFL ; which encompasses the parafacial respiratory group, pFRG) is a conditional oscillator that drives active expiration during periods of high respiratory demand, and increases ventilation through the recruitment of expiratory muscles. The pFL activity is highly modulated, and systematic analysis of its afferent projections is required to understand its connectivity and modulatory control. We combined a viral retrograde tracing approach to map direct brainstem projections to the putative location of pFL , with RNAScope and immunofluorescence to identify the neurochemical phenotype of the projecting neurons. Within the medulla, retrogradely-labeled, glutamatergic, glycinergic and GABAergic neurons were found in the ventral respiratory column (Bötzinger and preBötzinger Complex [preBötC], ventral respiratory group, ventral parafacial region [pFV ] and pFL ), nucleus of the solitary tract (NTS), reticular formation (RF), pontine and midbrain vestibular nuclei, and medullary raphe. In the pons and midbrain, retrogradely-labeled neurons of the same phenotypes were found in the Kölliker-Fuse and parabrachial nuclei, periaqueductal gray, pedunculopontine nucleus (PPT) and laterodorsal tegmentum (LDT). We also identified somatostatin-expressing neurons in the preBötC and PHOX2B immunopositive cells in the pFV , NTS, and part of the RF. Surprisingly, we found no catecholaminergic neurons in the NTS, A5 or Locus Coeruleus, no serotoninergic raphe neurons nor any cholinergic neurons in the PPT and LDT that projected to the pFL . Our results indicate that pFL neurons receive extensive excitatory and inhibitory inputs from several respiratory and nonrespiratory related brainstem regions that could contribute to the complex modulation of the conditional pFL oscillator for active expiration.

Transgene expression within the spinal cord of hTH-eGFP rats

The enzyme tyrosine hydroxylase (TH) is broadly expressed in catecholaminergic neurons. In the spinal cord, four components contain TH, including A11 diencephalospinal dopaminergic (DA-ergic) pathways, intraspinal DA-related neurons, supraspinal noradrenergic projections, and afferents of TH-expressing sensory neurons. A human TH-enhanced green fluorescent protein (hTH-eGFP) transgenic rat was recently developed to tag TH+ profiles in the nervous system for visualization. Using immunostaining, we found that only A11 pathways express GFP whereas the other 3 components do not in the spinal cord. Thus, this may suggest a genetic difference among these TH+ elements even though they produce the same protein.

Nuclear organization and morphology of catecholaminergic neurons and certain pallial terminal networks in the brain of the Nile crocodile, Crocodylus niloticus

In the current study, we use tyrosine hydroxylase (TH) immunohistochemistry to detail the nuclear parcellation and cellular morphology of neurons belonging to the catecholaminergic system in the brain of the Nile crocodile. In general, our results are similar to that found in another crocodilian (the spectacled caiman) and indeed other vertebrates, but certain differences of both evolutionary and functional significance were noted. TH immunopositive (TH+) neurons forming distinct nuclei were observed in the olfactory bulb (A16), hypothalamus (A11, A13-15), midbrain (A8-A10), pons (A5-A7) and medulla oblongata (area postrema, C1, C2, A1, A2), encompassing the more commonly observed nuclear complexes of this system across vertebrates. In addition, TH + neurons forming distinct nuclei not commonly identified in vertebrates were observed in the anterior olfactory nucleus, the pretectal nuclear complex, adjacent to the posterior commissure, and within nucleus laminaris, nucleus magnocellularis lateralis and the lateral vestibular nucleus. Palely stained TH + neurons were observed in some of the serotonergic nuclei, including the medial and lateral divisions of the superior raphe nucleus and the inferior raphe and inferior reticular nucleus, but not in other serotonergic nuclei. In birds, a high density of TH + fibres and pericellular baskets in the dorsal ventricular ridge marks the location of the nidopallium caudolaterale (NCL), a putative avian analogue of mammalian prefrontal cortex. In the dorsal ventricular ridge (DVR) of the crocodile a small region in the caudolateral anterior DVR (ADVRcl) revealed a slightly higher density of TH + fibres and some pericellular baskets (formed by only few TH + fibres). These results are discussed in an evolutionary and functional framework.

Selective deletion of Caspase-3 gene in the dopaminergic system exhibits autistic-like behaviour

Apoptotic caspases are thought to play critical roles in elimination of excessive and non-functional synapses and removal of extra cells during early developmental stages. Hence, an impairment of this process may thus constitute a basis for numerous neurological and psychiatric diseases. This view is especially relevant for dopamine due to its pleiotropic roles in motor control, motivation and reward processing. Here, we have analysed the effect of caspase-3 depletion on the development of catecholaminergic neurons and performed a wide array of neurochemical, ultrastructural and behavioural assays. To achieve this, we performed selective deletion of the Casp3 gene in tyrosine hydroxylase (TH)-expressing cells using Cre-loxP-mediated recombination. Histological evaluation of most relevant catecholaminergic nuclei revealed the ventral mesencephalon as the most affected region. Stereological analysis demonstrated an increase in the number of TH-positive neurons in both the substantia nigra and ventral tegmental area along with enlarged volume of the ventral midbrain. Analysis of main innervating tissues revealed a rather contrasting profile. In striatum, basal extracellular levels and potassium-evoked DA release were significantly reduced in mice lacking Casp3, a clear indication of dopaminergic hypofunction in dopaminergic innervating tissues. This view was sustained by analysis of TH-labelled dopaminergic terminals by confocal and electron microscopy. Remarkably, at a behavioural level, Casp3-deficient mice exhibited impaired social interaction, restrictive interests and repetitive stereotypies, which are considered the core symptoms of autism spectrum disorder (ASD). Our study revitalizes the potential involvement of dopaminergic transmission in ASD and provides an excellent model to get further insights in ASD pathogenesis.

Calbindin-D28K, parvalbumin, and calretinin in young and aged human locus coeruleus

Certain neuronal populations, including basal forebrain cholinergic neurons (BFCN) and noradrenergic neurons of the locus coeruleus (LC), are selectively vulnerable to pathology and loss early in the course of aging and Alzheimer's disease (AD). Human BFCN show substantial loss of the calcium-binding protein (CBP), calbindin-D28K (CB), during normal aging, which is associated with formation of neurofibrillary tangles and BFCN loss in AD. Here we determined if, similar to the BFCN, LC neurons contain CB or the other 2 ubiquitous CBPs parvalbumin and calretinin, and whether these proteins display an age-related loss from LC neurons. Immunostaining for CBP and tyrosine hydroxylase, a marker of catecholaminergic neurons, was used in sections from the LC of young and aged human brains. Parvalbumin and calretinin immunoreactivities were completely absent from human LC neurons. A subpopulation of LC neurons (~10%) contained CB immunoreactivity. Quantitative analysis revealed no age-related loss of CB from LC neurons. Thus, unlike the BFCN, age-related loss of CB does not figure prominently in the selective vulnerability of LC neurons to degeneration in AD.

“Sick-but-not-dead”: multiple paths to catecholamine deficiency in Lewy body diseases

Profound depletion of the catecholamines dopamine (DA) and norepinephrine in the brain, heart, or both characterizes Lewy body diseases such as Parkinson disease, dementia with Lewy bodies, and pure autonomic failure. Although one might presume that catecholamine deficiency in these disorders results directly and solely from loss of catecholaminergic neurons, there is increasing evidence that functional abnormalities in extant residual neurons contribute to the neurotransmitter deficiencies—the “sick-but-not-dead” phenomenon. This brief review highlights two such functional abnormalities—decreased vesicular sequestration of cytoplasmic catecholamines and decreased catecholamine biosynthesis. Another abnormality, decreased activity of aldehyde dehydrogenase, may have pathogenetic significance and contribute indirectly to the loss of catecholamine stores via interactions between the autotoxic catecholaldehyde 3,4-dihydroxyphenylacetaldehyde and the protein alpha-synuclein, which is a major component of Lewy bodies. Theoretically, chronically repeated stress responses could accelerate these abnormalities, via increased exocytosis and neuronal reuptake, which indirectly shifts tissue catecholamines from vesicular stores into the cytoplasm, and via increased tyrosine hydroxylation, which augments intra-cytoplasmic DA production. The discovery of specific paths mediating the sick-but-not-dead phenomenon offers novel targets for multi-pronged therapeutic approaches.

Noradrenergic But Not Dopaminergic Neurons Signal Task State Changes and Predict Reengagement After a Failure.

The two catecholamines, noradrenaline and dopamine, have been shown to play comparable roles in behavior. Both noradrenergic and dopaminergic neurons respond to cues predicting reward availability and novelty. However, even though both are thought to be involved in motivating actions, their roles in motivation have seldom been directly compared. We therefore examined the activity of putative noradrenergic neurons in the locus coeruleus and putative midbrain dopaminergic neurons in monkeys cued to perform effortful actions for rewards. The activity in both regions correlated with engagement with a presented option. By contrast, only noradrenaline neurons were also (i) predictive of engagement in a subsequent trial following a failure to engage and (ii) more strongly activated in nonrepeated trials, when cues indicated a new task condition. This suggests that while both catecholaminergic neurons are involved in promoting action, noradrenergic neurons are sensitive to task state changes, and their influence on behavior extends beyond the immediately rewarded action.

Brainstem catecholaminergic neurons mediate cognitive impairment in heart failure rats through regulation of hippocampal synaptic‐related gene expression.

Heart failure (HF) is common in elder population and it is highly associated with cognitive impairment (CI). Several studies suggest that autonomic imbalance contribute to age‐related CI. We previously showed that catecholaminergic neurons (C1) from the rostral ventrolateral medulla (RVLM) displayed chronic activation in HF. In addition, we also shown that HF rats showed CI. Whether RVLM‐C1 neurons contribute to CI in the setting of HF is completely unknown. Therefore, we aimed to study the contribution of RVLM‐C1 neurons on cognitive function in HF. Adult male Sprague‐Dawley rats underwent volume overload to induce HF. Anti‐dopamine β‐hydroxylase‐saporin (DβH–SAP: 5ng/150nl) was used to selectively destroy RVLM‐C1 neurons. Morris water maze (MWM) test was used to assess learning and memory performance. Micropunches containing the hippocampus were dissected to measure the expression of several synaptic‐related genes using a custom 12‐gene array by qRT‐PCR. HF rats displayed learning and memory impairment compared to control animals. Indeed, HF showed an increased traveled distance to the target platform along 5 trial/days during MWM test (15.1±0.5 vs. 30.7±2.1 m; Control vs. HF, respectively; p<.05). In addition, when the platform was changed daily to a different quadrant (i.e. to evaluate reference memory) HF rats showed an increase in the number of trials required to reach the platform compared to control rats (4.2±0.1 ± vs. 8.2±1.2 trials; Control vs. HF, respectively; p<.05). Partial ablation of RVLM‐C1 neurons in HF rats decreased traveled distance to reach the target platform compared to HF rats without treatment (18.1±1 vs 30.7±2.1 m; HFDβH–SAP vs. HF, respectively; p<.05). Also, during memory testing HF untreated rats needed almost twice the number of trials to accomplish the task when compared to HF+DβH–SAP rats (4.4±0.4 vs. 8.2±1.2 trials; HFDβH–SAP vs. HF, respectively; p<.05). Finally, compared to control rats, HF rats display (Ctrl vs. HF, in AU) decreased levels of PSD‐95 (1.0±0.2 vs. 0.4±0.3), Hommer1 (1.0±0.2 vs. 0.6±0.3), CaMKIV (1.0±0.2 vs. 0.5±0.3), Piccolo (1.0±0.3 vs. 0.4±0.2), Erc2 (1.0±0.3 vs. 0.2±0.1), RIM (1.0±0.5 vs. 0.3±0.2), Neuroligin 3 (1.0±0.4 vs. 0.5±0.3), Neuroxin 1 (1.0±0.3 vs. 0.6±0.4) and Synaptotagmin‐1 (1.0±0.3 vs. 0.6±0.1). Importantly, DβH–SAP treatment in HF significantly restored the expression levels of Hommer1, Piccolo, Erc2, RIM and Synaptotagmin‐1 to levels observed in control rats. Taken together, our results show that catecholaminergic RVLM neurons contribute to CI in HF rats. In addition, our results suggest that RVLM‐C1 may regulate hippocampal synaptic transmission in the setting of HF through the regulation of gene expression in the hippocampus.Support or Funding InformationSupported by FONDECYT 1180172 and 3190659 and the basal Center of Excellence in Aging and Regeneration (AFB 170005).

Selective optogenetic stimulation of RTN neurons drives ventilation and active expiration in unanesthetized rats

BackgroundThe retrotrapezoid nucleus (RTN) consists of neurons that are chemosensitive (i.e., CO2/pH sensitive), glutamatergic, Phox2b+ and Neuromedin B+ (Nmb+). Lentiviral vectors currently used to transduce RTN neurons with various opto‐ or pharmacogenetic actuators contain a Phox2b‐activated promoter, PRSx8. When injected into the RTN, these vectors also transduce substantial numbers of nearby C1 and catecholaminergic neurons. The purpose of the present study was to determine the effects on breathing elicited by selectively activating RTN neurons.MethodsTo selectively transduce RTN neurons we microinjected a mixture of Lenti‐PRSx8‐ChR2‐mCherry and AAV5‐FLEX‐taCasP3‐TEVp into the RTN of adult TH‐Cre rats. One month after the surgery, RTN neurons were stimulated with blue light (473 nm, 10 ms pulse, 20 Hz, for 20 s). All the measurements were conducted inside a plethysmography chamber. Diaphragm (DIA) and abdominal (ABD) muscle activity were recorded in conscious conditions. Results (mean ± SD), were analyzed using one‐ or two‐way ANOVA as required and differences were considered significant at P<0.05.ResultsRTN neurons (Nmb+, TH−) were selectively transduced (96.8 % ± 4) with the PRSx8 vector, the FLEX‐caspase AAV5 having caused the apoptotic death of C1 neurons (TH+) in the area where neurons were transduced with ChR2. Selective stimulation of RTN (NMB+ neurons; n=6) increased respiratory rate during wake and NREM sleep but not during REM sleep (Δ 30 ± 7 vs. 22.7 ± 6 vs. 4 ± 4 bpm from baseline), increased tidal volume also during quiet waking and non‐REM but not REM sleep (Δ 0.15 ± 0.07 vs. 0.13 ± 0.04 vs. 0.02 ± 0.007 mL/100g) and VE also during wake and non‐REM sleep but not REM (27.8 ± 11 vs. 21.4 ± 8 vs. 3.1 ±1 mL/100g/min). Selective RTN stimulation in rats breathing room air elicited active expiration (from 8.4 ± 7 to 15.9 ± 8 % of normalized abdominal EMG recorded at 9% FiCO2, n=3). RTN stimulation produced greater active expiration in rats exposed to 6% FiCO2 (from 56.3 ± 32 to 125.3 ± 104 % of normalized abdominal EMG).ConclusionsSelective RTN neuron stimulation drives breathing amplitude and frequency during wake, NREM but not in REM sleep, and elicits active expiration, especially at elevated levels of FiCO2.Support or Funding InformationSupported by grants from the National Institutes of Health (HL28785, 18 HL074011) and the Congenital Central Hypoventilation Syndrome Foundation to PGG.

MDMA and TAAR1‐mediated RhoA Activation in Serotonin Neurons

We have recently described the biochemical cascade through which the psychostimulant amphetamine leads to internalization of the dopamine and norepinephrine transporters, (DAT and NET). Amphetamine is a relatively high affinity substrate for DAT and NET and they both serve as efficient conduits for amphetamine entry into the cell. Once inside the cell, amphetamine activates the trace amine associated receptor 1 (TAAR1), a GPCR that we have shown couples to both GS and G13 signaling pathways. TAAR1 signaling through G13 increases the activity of RhoA, a small GTPase that regulates trafficking of the catecholamine carriers and increases their internalization. We wanted to investigate if similar G‐protein coupled signaling events also occur in serotonergic neurons. Although amphetamine does not have a particularly high affinity for the serotonin transporter (SERT), other TAAR1 ligands do, including 3,4‐methylenedioxymethamphetamine (MDMA).We developed primary cultures from the raphe nuclei of E15 mice. 3H‐serotonin uptake mediated by these cultures was largely citalopram sensitive, indicating SERT‐mediated serotonin transport. Pre‐treatment of these cultures with MDMA resulted in a decrease of SERT activity that could be blocked by an inhibitor of RhoA. Similarly, by total internal reflection fluorescence microscopy in HEK293 cells we found internalization of GFP‐tagged SERT in response to MDMA was blocked by a RhoA inhibitor. In HEK293 cells in which the TAAR1 gene has been deleted, MDMA did not trigger SERT internalization. In noradrenergic neurons cultured from the locus coeruleus or dopamine neurons from the midbrain, only higher doses of MDMA pretreatment could decrease transporter activity at the cell surface, likely reflecting the differences in affinity of the drug for the different monoamine transporters.These data indicate that pharmacological agents can regulate TAAR1 signaling in serotonergic, as well as catecholaminergic neurons and that the selectivity of TAAR1 agonists depends not only on affinity for the GPCR but also on their transport efficiency. Moreover, these findings suggest that the activation of TAAR1 in serotonin neurons and the subsequent internalization of surface SERT likely contribute to the modulation of serotonin signaling observed in response to MDMA.Support or Funding InformationThis work was supported by Intramural Research Program at the NIH, NIMH (ZIAMH002946).

Selective C1 neuron stimulation restores blood pressure (BP) and heart rate (HR) during decompensated hemorrhage in unanesthetized rats

BackgroundDuring hemorrhage, BP is initially maintained near normal (compensated phase) by an increase in sympathetic nerve activity (SNA). However, further blood loss (>20–25% blood volume) causes an abrupt and severe BP drop (late or “decompensated” phase) associated with an unexplained reduction of SNA and HR. Under several circumstances (e.g. hypoxia, anesthesia) BP is largely determined by the level of activity of catecholaminergic presympathetic neurons (C1 neurons) located within the rostral ventrolateral medulla (RVLM).AimIn this study we sought to determine the contribution of the C1 neurons to BP and HR during the compensated and decompensated phases of hemorrhage in non‐anesthetized rats. Additionally, we tested whether C1 neuron stimulation can restore BP and HR during the decompensated phase.MethodsWe determined the contribution of the C1 neurons to mean arterial pressure (MAP) and HR by measuring how much these variables change when these neurons are selectively silenced or activated using loss‐ or gain‐of‐function optogenetics. To inhibit C1 neurons, they were transduced with an inhibitory opsin (Archaerhodopsin) by microinjecting a Cre‐dependent vector (AAV2‐DIO‐ArchT3.0‐eYFP) bilaterally into the RVLM of male and female adult TH‐Cre rats. To stimulate C1 neurons, they were transduced with an excitatory opsin (Channelrhodopsin, AAV2‐DIO‐ChR2‐eYFP) as described above. Four weeks later, C1 neurons were inhibited by delivering green light (532 nm, continuous pulse for 10 s) or stimulated by delivering blue light (473 nm, 5 ms pulse, 10 Hz, for 10 s) at regular intervals while the rats were subjected to a controlled hypotensive hemorrhage (18 ml/kg of blood withdrawal over 20 minutes, rate 0.3–0.5 ml/min). Data are expressed as mean ± SD and were analyzed using one‐way ANOVA for repeated measures. Differences were considered significant when p<0.05.ResultsDecompensated phase of hemorrhage was marked by a decrease in MAP (from 101 ± 8 to 53 ± 10 mmHg) and bradycardia (from 326 ± 20 to 245 ± 44 bpm). Bilateral C1 neurons inhibition produced a larger MAP reduction during the early compensated phase of hemorrhage than at rest (Δ MAP: −6 ± 2 vs. −23 ± 5 mmHg, n=9). However, the MAP drop elicited by C1 cell inhibition during the decompensated phase of hemorrhage (Δ MAP: −4 ± 4 mmHg) was no different from the drop at rest. Bilateral C1 neuron stimulation increased BP at rest (Δ MAP: 35 ± 6 mmHg) and raised BP to pre‐hemorrhage levels during the decompensated phase of hemorrhage (Δ MAP: 42 ± 5 mmHg, n=6). C1 stimulation, decreased HR at rest whereas it increased HR during the decompensated hemorrhage (Δ HR: −14 ± 36 vs. +58 ± 22 bpm).ConclusionsC1 neuron activation contributes to BP stability during the early compensated phase of hemorrhage. C1 neurons are probably inhibited during the decompensated phase of hemorrhage. Their re‐activation rescues BP and HR, indicating that these neurons remain potentially capable of maintaining BP despite the blood loss.Support or Funding InformationNational Institutes of Health, RO1 (HL028785) to P.G.G. and American Heart Association (19POST34430205) to G.M.P.R.S.

Hypocretin (orexin) immunoreactivity in the feline midbrain: Relevance for the generation of wakefulness

Hypocretins (Hcrt) 1 and 2 are two neuropeptides synthesized from neurons that are located in the perifornical area of the lateral hypothalamus. These neurons project diffusely throughout the central nervous system, and have been implicated in the generation and maintenance of wakefulness, as well as in critical physiological processes that occur during this behavioral state, such as motivation.The hypocretinergic projections towards the feline midbrain have not been studied before. Therefore, the aim of the present study was to analyze their relationship to the midbrain neurons, that are critically involved in the control of sleep and wakefulness. With this purpose, we examined the distribution of Hcrt1-positive fibers in the midbrain and pontomesencephalic area of the domestic cat (Felis catus), and their relationship with catecholaminergic and cholinergic neurons by means of single and double immunohistochemistry. Hcrtergic axons with distinctive varicosities and buttons were heterogeneously distributed, exhibiting different densities in distinct regions of the midbrain. High Hcrtergic fiber densities were observed in the periaqueductal gray, interpeduncular nucleus, locus coeruleus and cholinergic mesopontine regions. In addition, we studied in detail the Hcrtergic projection towards the dopaminergic nuclei of the midbrain. While very few Hcrt + fibers were observed in the substantia nigra pars compacta, the highest density of Hcrtergic fibers was found in the dopaminergic ventral periaqueductal gray area (also called A10dc area); appositions between Hcrtergic terminals and dopaminergic somata and dendrites were observed within this area. Because this dopaminergic area has been involved in the control of wakefulness, the present anatomical data provides relevant support about the role of the Hcrtergic system in the generation of this behavioral state.

Effects on goal directed behavior and habit in two animal models of Parkinson's disease.

Instrumental conditioning involves two different processes: Goal-directed behavior, characterized by its dependence on the causal relationship between action and outcome and the sensitivity of actions to changes in the value of the outcome; and habits, characterized for its persistence and insensitivity to changes after conditioning. It is known that the dopaminergic system is involved in both kind of learning. The present experiments analyzed two animal models of Parkinson's disease. The 6-OHDA model causes selective damage of the catecholaminergic neurons, specifically affecting the dopaminergic neurons in nigro-striatal system. This model simulates degenerative process symptomatology of Parkinson's disease. On the other hand, the LPS model generates an inflammation process in the infusion area. This model simulates the early symptoms of this disorder, including neuroinflammation and microglia activation. In order to validate both parkinsonian models, we studied if 6-OHDA and LPS models cause the same behavioral effects. The results showed that the 6-OHDA model interfered with the process involved in habit formation. In contrast, animals treated with LPS showed a goal-directed learning deficit. Differences between these models could be due to the different effects on Substantia Nigra neurons. 6-OHDA model might disrupt the nigrostriatal pathway, while LPS could interfere on efferences and afferences to Substantia Nigra.

Distributions of hypothalamic neuron populations coexpressing tyrosine hydroxylase and the vesicular GABA transporter in the mouse.

The hypothalamus contains catecholaminergic neurons marked by the expression of tyrosine hydroxylase (TH). As multiple chemical messengers coexist in each neuron, we determined if hypothalamic TH-immunoreactive (ir) neurons express vesicular glutamate or GABA transporters. We used Cre/loxP recombination to express enhanced GFP (EGFP) in neurons expressing the vesicular glutamate (vGLUT2) or GABA transporter (vGAT), then determined whether TH-ir neurons colocalized with native EGFPVglut2 - or EGFPVgat -fluorescence, respectively. EGFPVglut2 neurons were not TH-ir. However, discrete TH-ir signals colocalized with EGFPVgat neurons, which we validated by in situ hybridization for Vgat mRNA. To contextualize the observed pattern of colocalization between TH-ir and EGFPVgat , we first performed Nissl-based parcellation and plane-of-section analysis, and then mapped the distribution of TH-ir EGFPVgat neurons onto atlas templates from the Allen Reference Atlas (ARA) for the mouse brain. TH-ir EGFPVgat neurons were distributed throughout the rostrocaudal extent of the hypothalamus. Within the ARA ontology of gray matter regions, TH-ir neurons localized primarily to the periventricular hypothalamic zone, periventricular hypothalamic region, and lateral hypothalamic zone. There was a strong presence of EGFPVgat fluorescence in TH-ir neurons across all brain regions, but the most striking colocalization was found in a circumscribed portion of the zona incerta (ZI)-a region assigned to the hypothalamus in the ARA-where every TH-ir neuron expressed EGFPVgat . Neurochemical characterization of these ZI neurons revealed that they display immunoreactivity for dopamine but not dopamine β-hydroxylase. Collectively, these findings indicate the existence of a novel mouse hypothalamic population that may signal through the release of GABA and/or dopamine.

Heart to breathe: partial ablation of rostral ventrolateral medulla catecholaminergic neurons mediates disordered breathing in volume overload heart failure rats

Heart to breathe: partial ablation of rostral ventrolateral medulla catecholaminergic neurons mediates disordered breathing in volume overload heart failure rats

Role of Glial Cell Line-Derived Neurotrophic Factor in the Maintenance of Adult Mesencephalic Catecholaminergic Neurons.

The glial cell line-derived neurotrophic factor has a potent neuroprotective action on mesencephalic dopamine neurons, which are progressively lost in Parkinson's disease. Intrastriatal administration of this factor is a promising therapy for Parkinson's disease. Glial cell line-derived neurotrophic factor is naturally produced in restricted cerebral regions, such as the striatum, septum, and thalamus; however, its effects in the adult brain remain under debate. We sought to clarify the physiologic role of endogenous glial cell line-derived neurotrophic factor in the survival of catecholaminergic neurons of the substantia nigra pars compacta and the locus coeruleus in adult mice. We used 2 new Cre recombinase-based mouse models to delete a floxed-glial cell line-derived neurotrophic factor gene. The first model had Cre expression in the parvalbumin expressing interneurons, as these cells represent the major source of striatal glial cell line-derived neurotrophic factor. The second model was an estrogen receptor 2-based inducible Cre triggered by tamoxifen at 2 months of age. We found that the floxed-glial cell line-derived neurotrophic factor gene was resilient to ablation by Cre-induced recombination and that parvalbumin-driven Cre was particularly inefficient to do so. The inducible-Cre model allowed an average 70% to 80% reduction in glial cell line-derived neurotrophic factor messenger ribonucleic acid and protein in striatum and septum with moderate significant loss of catecholamine neurons in the nigrostriatal pathway and, more markedly, in the locus coeruleus. This was accompanied with mild locomotor decline. Our data support qualitatively the view that brain glial cell line-derived neurotrophic factor is needed for the maintenance of adult central catecholaminergic neurons. © 2020 International Parkinson and Movement Disorder Society.

Исследование иннервации двенадцатиперстной кишки крысы с использованием нейральных иммуногистохимических маркеров

The purpose of the work is to study the topography, morphology, and mediator status of the nerve apparatus of the rat duodenum using immunohistochemical methods. The use of markers selective for the peripheral nervous system (PGP 9.5, synaptophysin (Syn), serotonin (5HT) and tyrosine hydroxylase (TH) made it possible to reveal in the tissues of the intestinal wall, in addition to the two main submucosal and myienteral ganglionic nerve plexuses), the third plexus is the mucous epithelium villous. Using comparative immunohistochemical analysis, it was found that most of the nerve structures (neurons, nerve fibers, interneuronal and neuromuscular synapses in the ganglion plexuses) are PGP 9.5-immunopositive and cholinergic. There are no catecholaminergic and serotonergic (5HT +) neurons in the studied ganglion plexuses. It is assumed that postganglionic sympathetic fibers enter the duodenum from the outside. They participate in the innervation of the muscle layers of the intestinal wall and arterial vessels, form pericellular synapses on the bodies and dendrites of parasympathetic neurons. A large number of terminal varicose axons were found in the connective tissue of the villi using PGP 9.5, and single bipolar neuron-like cells were found in the epithelium. Sympathetic elements in the villi were not identified. A reaction to serotonin revealed a large number of serotonin-containing cells that were morphologically similar to enterochromaffinocytes.

Development of early diagnosis of Parkinson's disease based on the search for biomarkers such as premotor symptoms and changes in blood

To determine changes in the chemical composition of blood plasma in subjects at risk of Parkinson's disease (PD) at the prodromal stage compared with age control. Subjects at risk were selected for the presence of characteristic premotor symptoms, including impairments of sleep, olfaction and constipation.The risk group included 12 people, the control group - 8 people. Among seven catecholamines and their metabolites detected in the blood, only the concentration of L-dioxiphenylalanine (L-DOPA) changed (decreased) in subjects at risk compared with the control. A decrease in the concentration of L-DOPA is considered as a manifestation (marker) of selective degeneration of central and peripheral catecholaminergic neurons in PD. In contrast to L-DOPA, the concentration of seven of the twelve detected sphingomyelins in the blood of the subjects at risk increased. Given that a change in the metabolism of sphingomyelins is associated with processes such as apoptosis, autophagy, and synucleinopathy, an increase in their concentration in the blood of patients at risk is considered as a manifestation of systemic general degeneration of central and peripheral neurons. Finally, in the blood of subjects at risk, we found a trend towards a decrease in the concentration of urates, which are endogenous neuroprotectors. The changes in the level of L-DOPA, sphingmyelins and urates in the blood of subjects at risk may serve as diagnostic markers of PD at the prodromal stage.

Rostral ventrolateral medullary catecholaminergic neurones mediate irregular breathing pattern in volume overload heart failure rats.

A strong association between disordered breathing patterns, elevated sympathetic activity, and enhanced central chemoreflex drive has been shown in experimental and human heart failure (HF). The aim of this study was to determine the contribution of catecholaminergic rostral ventrolateral medulla catecholaminergic neurones (RVLM-C1) to both haemodynamic and respiratory alterations in HF. Apnoea/hypopnoea incidence (AHI), breathing variability, respiratory-cardiovascular coupling, cardiac autonomic control and cardiac function were analysed in HF rats with or without selective ablation of RVLM-C1 neurones. Partial lesion (∼65%) of RVLM-C1 neurones reduces AHI, respiratory variability, and respiratory-cardiovascular coupling in HF rats. In addition, the deleterious effects of central chemoreflex activation on cardiac autonomic balance and cardiac function in HF rats was abolished by ablation of RVLM-C1 neurones. Our findings suggest that RVLM-C1 neurones play a pivotal role in breathing irregularities in volume overload HF, and mediate the sympathetic responses induced by acute central chemoreflex activation. Rostral ventrolateral medulla catecholaminergic neurones (RVLM-C1) modulate sympathetic outflow and breathing under normal conditions. Heart failure (HF) is characterized by chronic RVLM-C1 activation, increased sympathetic activity and irregular breathing patterns. Despite studies showing a relationship between RVLM-C1 and sympathetic activity in HF, no studies have addressed a potential contribution of RVLM-C1 neurones to irregular breathing in this context. Thus, the aim of this study was to determine the contribution of RVLM-C1 neurones to irregular breathing patterns in HF. Sprague-Dawley rats underwent surgery to induce volume overload HF. Anti-dopamine β-hydroxylase-saporin toxin (DβH-SAP) was used to selectively lesion RVLM-C1 neurones. At 8weeks post-HF induction, breathing pattern, blood pressures (BP), respiratory-cardiovascular coupling (RCC), central chemoreflex function, cardiac autonomic control and cardiac function were studied. Reduction (∼65%) of RVLM-C1 neurones resulted in attenuation of irregular breathing, decreased apnoea-hypopnoea incidence (11.1±2.9vs. 6.5±2.5 eventsh-1 ; HF+Veh vs. HF+DβH-SAP; P<0.05) and improved cardiac autonomic control in HF rats. Pathological RCC was observed in HF rats (peak coherence >0.5 between breathing and cardiovascular signals) and was attenuated by DβH-SAP treatment (coherence: 0.74±0.12vs. 0.54±0.10, HF+Veh vs. HF+DβH-SAP rats; P<0.05). Central chemoreflex activation had deleterious effects on cardiac function and cardiac autonomic control in HF rats that were abolished by lesion of RVLM-C1 neurones. Our findings reveal that RVLM-C1 neurones play a major role in irregular breathing patterns observed in volume overload HF and highlight their contribution to cardiac dysautonomia and deterioration of cardiac function during chemoreflex activation.

A membrane-bound dopamine β-hydroxylase highly expressed in granulocyte of Pacific oyster Crassostrea gigas

Dopamine β-hydroxylase (DBH) is one of key rate-limiting enzymes converting dopamine to norepinephrine. It locates not only in catecholaminergic neuron system, but also in immunocytes and plays roles in the immune response of vertebrates. However, the knowledge about the function of DBH in immune system is still very limited in invertebrates. In the present study, the DBH gene family with seven members was screened from Crassostrea gigas genome, and their mRNA expressions in various tissues were recorded. Among them, one DBH (designated CgDBH-1) with high expression level in oyster hemocytes was further characterized. The deduced amino acid sequence of CgDBH-1 was predicted to contain a transmembrane domain and shared 30.1% and 30.9% similarity with that in Mus musculus and Homo sapiens, respectively. CgDBH-1 was closely clustered with DBH from Aplysia californica in the phylogenetic tree. The recombinant protein of CgDBH-1 (rCgDBH-1) exhibited significant enzymatic activity (0.54 ± 0.019 pmol L−1 min−1) to synthesize norepinephrine. Importantly, the mRNA transcript of CgDBH-1 was highly expressed in oyster hemocytes, and the highest expression level was observed in granulocytes among the three types of hemocytes, which was 8.18-fold (p < 0.01) of that in agranulocytes. Moreover, the expression of CgDBH-1 in hemocytes was significantly increased at the late stage of immune response. The CgDBH-1 protein was mainly co-localized with the granules and endoplasmic reticulum (ER) of granulocytes. These results collectively suggested that CgDBH-1, as a novel molluscan norepinephrine synthesizing enzyme highly expressed in granulocytes, involved in the late-stage immune response of oysters, which provided vital insight to understand the crosstalk between neuroendocrine and immune systems in invertebrates.