Subcellular and regional localization of mRNA translation in midbrain dopamine neurons

Local translation within excitatory and inhibitory neurons is involved in neuronal development and synaptic plasticity. Despite the extensive dendritic and axonal arborizations of central monoaminergic neurons, the subcellular localization of protein synthesis is not well-characterized in these populations. Here, we investigated mRNA localization and translation in midbrain dopaminergic (mDA) neurons, cells with enormous axonal and dendritic projections, both of which exhibit stimulation-evoked dopamine (DA) release. Using highly-sensitive ribosome-bound RNA-sequencing and imaging approaches in mDA axons, we found no evidence for axonal mRNA localization or translation. In contrast, mDA neuronal dendritic projections into the substantia nigra reticulata (SNr) contain ribosomes and mRNAs encoding the major components of DA synthesis, release, and reuptake machinery. Surprisingly, we also observed dendritic localization of mRNAs encoding synaptic vesicle-related proteins, including those involved in vesicular exocytic fusion. Our results are consistent with a role for local translation in the regulation of DA release from dendrites, but not from axons. Our translatome data further defined a molecular signature of the sparse mDA neurons resident in the SNr, including enrichment of Atp2a3/SERCA3, an ER calcium pump previously undescribed in mDA neurons.

Stress decreases serotonin tone in the nucleus accumbens to promote aversion and potentiate cocaine preference via decreased stimulation of 5-HT1B receptors

Stress-induced release of dynorphins (Dyn) activates kappa opioid receptors (KOR) in monoaminergic neurons to produce dysphoria and potentiate drug reward; however, the circuit mechanisms responsible for this effect are not known. We found that conditional deletion of KOR from Slc6a4 (SERT)-expressing neurons blocked stress-induced potentiation of cocaine conditioned place preference (CPP). Within the dorsal raphe nucleus (DRN), two overlapping populations of KOR-expressing neurons: Slc17a8 (VGluT3) and SERT, were distinguished functionally and anatomically. Optogenetic inhibition of these SERT+ neurons potentiated subsequent cocaine CPP, whereas optical inhibition of the VGluT3+ neurons blocked subsequent cocaine CPP. SERT+/VGluT3- expressing neurons were concentrated in the lateral aspect of the DRN. SERT projections from the DRN were observed in the medial nucleus accumbens (mNAc), but VGluT3 projections were not. Optical inhibition of SERT+ neurons produced place aversion, whereas optical stimulation of SERT+ terminals in the mNAc attenuated stress-induced increases in forced swim immobility and subsequent cocaine CPP. KOR neurons projecting to mNAc were confined to the lateral aspect of the DRN, and the principal source of dynorphinergic (Pdyn) afferents in the mNAc was from local neurons. Excision of Pdyn from the mNAc blocked stress-potentiation of cocaine CPP. Prior studies suggested that stress-induced dynorphin release within the mNAc activates KOR to potentiate cocaine preference by a reduction in 5-HT tone. Consistent with this hypothesis, a transient pharmacological blockade of mNAc 5-HT1B receptors potentiated subsequent cocaine CPP. 5-HT1B is known to be expressed on 5-HT terminals in NAc, and 5-HT1B transcript was also detected in Pdyn+, Adora2a+ and ChAT+ (markers for direct pathway, indirect pathway, and cholinergic interneurons, respectively). Following stress exposure, 5-HT1B transcript was selectively elevated in Pdyn+ cells of the mNAc. These findings suggest that Dyn/KOR regulates serotonin activation of 5HT1B receptors within the mNAc and dynamically controls stress response, affect, and drug reward.

INTEROCEPTIVE INFORMATION OF PHYSICAL VIGOR: OREXIN NEURONS GAUGE CIRCULATING IGF-I FOR MOTIVATIONAL MOTOR OUTPUT

Brain regulation of bodily functions requires interoceptive feedback signals carrying information about the periphery. As mice with low serum IGF-I levels (LID mice) show reduced spontaneous physical activity, we speculated that body vigor information might be conveyed by circulating IGF-I, a regulator of skeletal muscle and bone mass that enters the brain during physical activity. Since hypothalamic orexin neurons, that are involved in regulating physical activity, express IGF-I receptors (IGF-IR), we hypothesized that these neurons might gauge circulating IGF-I levels. Inactivation of IGF-IR in mouse orexin neurons (Firoc mice) reduced spontaneous activity. Firoc mice maintain normal physical fitness but show anxiety- and depressive-like behaviors that seems to interfere with the rewarding effects of exercise, as they were less sensitive to the rewarding actions of exercise. Further, in response to exercise, Firoc mice showed reduced activation of hypothalamic orexin neurons and ventro-tegmental area (VTA) monoaminergic neurons, as indicated by c-fos staining. Collectively, these results suggest that circulating IGF-I is gauged by orexin neurons to modulate physical activity in part by stimulation of the VTA to motivate motor output. Hence, serum IGF-I may constitute a feedback signal, informing orexin neurons to adapt physical activity to physical vigor.

Neuropathology of the Brainstem to Mechanistically Understand and to Treat Alzheimer’s Disease

Alzheimer’s disease (AD) is a devastating neurodegenerative disorder as yet without effective therapy. Symptoms of this disorder typically reflect cortical malfunction with local neurohistopathology, which biased investigators to search for focal triggers and molecular mechanisms. Cortex, however, receives massive afferents from caudal brain structures, which do not only convey specific information but powerfully tune ensemble activity. Moreover, there is evidence that the start of AD is subcortical. The brainstem harbors monoamine systems, which establish a dense innervation in both allo- and neocortex. Monoaminergic synapses can co-release neuropeptides either by precisely terminating on cortical neurons or, when being “en passant”, can instigate local volume transmission. Especially due to its early damage, malfunction of the ascending monoaminergic system emerges as an early sign and possible trigger of AD. This review summarizes the involvement and cascaded impairment of brainstem monoaminergic neurons in AD and discusses cellular mechanisms that lead to their dysfunction. We highlight the significance and therapeutic challenges of transmitter co-release in ascending activating system, describe the role and changes of local connections and distant afferents of brainstem nuclei in AD, and summon the rapidly increasing diagnostic window during the last few years.

Sleep, Narcolepsy and Sodium Oxybate

: Sodium oxybate (SO) has been in use for many decades to treat narcolepsy with cataplexy. It functions as a weak GABAB agonist but also as an energy source for the brain as a result of its metabolism to succinate and as a powerful antioxidant because of its capacity to induce the formation of NADPH. Its actions at thalamic GABAB receptors can induce slow wave activity while its actions at GABAB receptors on monoaminergic neurons can induce or delay REM sleep. By altering the balance between monoaminergic and cholinergic neuronal activity, SO uniquely can induce and prevent cataplexy. The formation of NADPH may enhance sleep’s restorative process by accelerating the removal of the reactive oxygen species (ROS) which accumulate during wakefulness. SO improves alertness in patients with narcolepsy and in normal subjects. SO may allay severe psychological stress - an inflammatory state triggered by increased levels of ROS and characterized by cholinergic supersensitivity and monoaminergic deficiency. SO may be able to eliminate the inflammatory state and correct the cholinergic/ monoaminergic imbalance.

Stress induces divergent gene expression among lateral habenula efferent pathways

The lateral habenula (LHb) integrates critical information regarding aversive stimuli that shapes decision making and behavioral responses. The three major LHb outputs innervate dorsal raphe nucleus (DRN), ventral tegmental area (VTA), and the rostromedial tegmental nucleus (RMTg). LHb neurons that project to these targets are segregated and nonoverlapping, and this led us to consider whether they have distinct molecular phenotypes and adaptations to stress exposure. In order to capture a time-locked profile of gene expression after repeated forced swim stress, we used intersectional expression of RiboTag in rat LHb neurons and next-gen RNA sequencing to interrogate the RNAs actively undergoing translation from each of these pathways. The “translatome” in the neurons comprising these pathways was similar at baseline, but diverged after stress, especially in the neurons projecting to the RMTg. Using weighted gene co-expression network analysis, we found one module comprising genes downregulated after stress in the RMTg-projecting LHb neurons; there was an overrepresentation of genes associated with phosphoinositide 3 kinase (PI3K) signaling in this module. Reduced PI3K signaling in RMTg-projecting LHb neurons may be a compensatory adaptation that alters the functional balance of LHb outputs to GABAergic vs. monoaminergic neurons following repeated stress exposure.

L-DOPA in Parkinson’s Disease: Looking at the “False” Neurotransmitters and Their Meaning

L-3,4-dihydroxyphenylalanine (L-DOPA) has been successfully used in the treatment of Parkinson’s disease (PD) for more than 50 years. It fulfilled the criteria to cross the blood–brain barrier and counteract the biochemical defect of dopamine (DA). It remarkably worked after some adjustments in line with the initial hypothesis, leaving a poor place to the plethora of mechanisms involving other neurotransmitters or mechanisms of action beyond newly synthesized DA itself. Yet, its mechanism of action is far from clear. It involves numerous distinct cell populations and does not mimic the mechanism of action of dopaminergic agonists. L-DOPA-derived DA is mainly released by serotonergic neurons as a false neurotransmitter, and serotonergic neurons are involved in L-DOPA-induced dyskinesia. The brain pattern and magnitude of DA extracellular levels together with this status of false neurotransmitters suggest that the striatal effects of DA via this mechanism would be minimal. Other metabolic products coming from newly formed DA or through the metabolism of L-DOPA itself could be involved. These compounds can be trace amines and derivatives. They could accumulate within the terminals of the remaining monoaminergic neurons. These “false neurotransmitters,” also known for some of them as inducing an “amphetamine-like” mechanism, could reduce the content of biogenic amines in terminals of monoaminergic neurons, thereby impairing the exocytotic process of monoamines including L-DOPA-induced DA extracellular outflow. The aim of this review is to present the mechanism of action of L-DOPA with a specific attention to “false neurotransmission.”

LIPOPOLYSACCHARIDE-INDUCED IMMUNOLOGICAL STRESS AT EARLY STAGES OF PREGNANCY AFFECTS THE DEVELOPMENT OF GONADOTROPIN RELEASING-HORMONE (GNRH)-PRODUCING SYSTEM

The aim of the present work was to study the development of afferent bonds between GnRH- and monoaminergic neurons in rat fetuses and to identify possible targets affected by LPS-induced inflammation. The innervation was analyzed using retrograde tracing method with DiI dye. At ED17 and ED21 olfactory bulbs (the area of GnRH migration) are innervated with monoaminergic neurons of septum and in lateral hypothalamus. The GnRH- and monoaminergic neuron interaction zones are sensitive to LPS (E. coli) prenatal exposure, which induces pro-inflammatory cytokine synthesis. We suppose that the olfactory bulbs of fetal forebrain can be a possible area of cytokine influence on GnRH- and monoaminergic neuron interaction.

Descending monoaminergic pathways projecting to the spinal defecation center enhance colorectal motility in rats

The central regulating mechanisms of defecation, especially roles of the spinal defecation center, are still unclear. We have shown that monoamines including norepinephrine, dopamine, and serotonin injected into the spinal defecation center cause propulsive contractions of the colorectum. These monoamines are the main neurotransmitters of descending pain inhibitory pathways. Therefore, we hypothesized that noxious stimuli in the colorectum would activate the descending monoaminergic pathways projecting to the spinal defecation center and that subsequently released endogenous monoamine neurotransmitters would enhance colorectal motility. Colorectal motility was measured in rats anesthetized with α-chloralose and ketamine. As a noxious stimulus, capsaicin was administered into the colorectal lumen. To interrupt neuronal transmission in the spinal defecation center, antagonists of norepinephrine, dopamine, and/or serotonin receptors were injected intrathecally at the L6-S1 spinal level, where the spinal defecation center is located. Intraluminal administration of capsaicin, acting on the transient receptor potential vanilloid 1 channel, caused transient propulsive contractions. The effect of capsaicin was abolished by surgical severing of the pelvic nerves or thoracic spinal transection at the T4 level. Capsaicin-induced contractions were blocked by preinjection of D2-like dopamine receptor and 5-hydroxytryptamine subtype 2 and 3 receptor antagonists into the spinal defecation center. We demonstrated that intraluminally administered capsaicin causes propulsive colorectal motility through reflex pathways involving the spinal and supraspinal defecation centers. Our results provide evidence that descending monoaminergic neurons are activated by noxious stimulation to the colorectum, leading to facilitation of colorectal motility. NEW & NOTEWORTHY The present study demonstrates that noxious stimuli in the colorectum activates the descending monoaminergic pathways projecting to the spinal defecation center and that subsequently released endogenous monoamine neurotransmitters, serotonin and dopamine, enhance colorectal motility. Our findings provide a possible explanation of the concurrent appearance of abdominal pain and bowel disorder in irritable bowel syndrome patients. Thus the present study may provide new insights into understanding of mechanisms of colorectal dysfunction involving the central nervous system.