An excitatory peri-tegmental reticular nucleus circuit for wake maintenance.

Sleep is a necessity for our survival, but its regulation remains incompletely understood. Here, we used a human sleep duration gene to identify a population of cells in the peri-tegmental reticular nucleus (pTRNADRB1) that regulate sleep-wake, uncovering a role for a poorly understood brain area. Although initial ablation in mice led to increased wakefulness, further validation revealed that pTRNADRB1 neuron stimulation strongly promotes wakefulness, even after stimulation offset. Using combinatorial genetics, we found that excitatory pTRNADRB1 neurons promote wakefulness. pTRN neurons can be characterized as anterior- or posterior-projecting neurons based on multiplexed analysis of projections by sequencing (MAPseq) analysis. Finally, we found that pTRNADRB1 neurons promote wakefulness, in part, through projections to the lateral hypothalamus. Thus, human genetic information from a human sleep trait allowed us to identify a role for the pTRN in sleep-wake regulation.

The ultrastructure of macaque mesencephalic trigeminal nucleus neurons.

Primary afferents originating from the mesencephalic trigeminal nucleus provide the main source of proprioceptive information guiding mastication, and thus represent an important component of this critical function. Unlike those of other primary afferents, their cell bodies lie within the central nervous system. It is believed that this unusual central location allows them to be regulated by synaptic input. In this study, we explored the ultrastructure of macaque mesencephalic trigeminal nucleus neurons to determine the presence and nature of this synaptic input in a primate. We first confirmed the location of macaque mesencephalic trigeminal neurons by retrograde labeling from the masticatory muscles. Since the labeled neurons were by far the largest cells located at the edge of the periaqueductal gray, we could undertake sampling for electron microscopy based on soma size. Ultrastructurally, mesencephalic trigeminal neurons had very large somata with euchromatic nuclei that sometimes displayed deeply indented nuclear membranes. Terminal profiles with varied vesicle characteristics and synaptic density thicknesses were found in contact with either their somatic plasma membranes or somatic spines. However, in contradistinction to other, much smaller, somata in the region, the plasma membranes of the mesencephalic trigeminal somata had only a few synaptic contacts. They did extend numerous somatic spines of various lengths into the neuropil, but most of these also lacked synaptic contact. The observed ultrastructural organization indicates that macaque trigeminal mesencephalic neurons do receive synaptic contacts, but despite their central location, they only avail themselves of very limited input.

Subsystem macroarchitecture of the intrinsic midbrain neural network and its tectal and tegmental subnetworks.

The midbrain is the smallest of three primary vertebrate brain divisions. Here we use network science tools to reveal the global organizing principles of intramidbrain axonal circuitry before adding extrinsic connections with the remaining nervous system. Curating the experimental neuroanatomical literature yielded 17,248 connection reports for 8,742 possible connections between the 94 gray matter regions forming the right and left midbrain. Evidence for the existence of 1,676 connections suggests a 19.2% connection density for this network, similar to that for the intraforebrain network [L. W. Swanson et al., Proc. Natl. Acad. Sci. U.S.A. 117, 31470-31481 (2020)]. Multiresolution consensus cluster analysis parceled this network into a hierarchy with 6 top-level and 30 bottom-level subsystems. A structure-function model of the hierarchy identifies midbrain subsystems that play specific functional roles in sensory-motor mechanisms, motivation and reward, regulating complex reproductive and agonistic behaviors, and behavioral state control. The intramidbrain network also contains four bilateral region pairs designated putative hubs. One pair contains the superior colliculi of the tectum, well known for participation in visual sensory-motor mechanisms, and the other three pairs form spatially compact right and left units (the ventral tegmental area, retrorubral area, and midbrain reticular nucleus) in the tegmentum that are implicated in motivation and reward mechanisms. Based on the core hypothesis that subsystems form functionally cohesive units, the results provide a theoretical framework for hypothesis-driven experimental analysis of neural circuit mechanisms underlying behavioral responses mediated in part by the midbrain.

Cerebellar Inputs in the American Alligator (Alligator mississippiensis).

Crocodilians (alligators, crocodiles, and gharials) are the closet living relatives to birds and, as such, represent a key clade to understand the evolution of the avian brain. However, many aspects of crocodilian neurobiology remain unknown. In this paper, we address an important knowledge gap as there are no published studies of cerebellar connections in any crocodilian species. We used injections of retrograde tracers into the cerebellum of the American alligator (Alligator mississippiensis) to describe for the first time the origin of climbing and mossy fiber inputs. We found that inputs to the cerebellum in the American alligator are similar to those of other nonavian reptiles and birds. Retrograde labeled cells were found in the spinal cord, inferior olive, reticular formation, vestibular and cerebellar nuclei, as well as in nucleus ruber and surrounding tegmentum. Additionally, we found no retrogradely labeled cells in the anterior rhombencephalon which suggest that, like other nonavian reptiles, crocodilians may lack pontine nuclei. Similar to birds and other nonavian reptiles, we found inputs to the cerebellum from the pretectal nucleus lentiformis mesencephali. Additionally, we found retrogradely labeled neurons in two nuclei in the pretectum: the nucleus circularis and the interstitial nucleus of the posterior commissure. These pretectal projections have not been described in any other nonavian reptile to date, but they do resemble projections from the nucleus spiriformis medialis of birds. Our results show that many inputs to the cerebellum are highly conserved among sauropsids and that extensive pretectal inputs to the cerebellum are not exclusive to the avian brain. Finally, we suggest that the pontine nuclei of birds are an evolutionary novelty that may have evolved after the last common ancestor between birds and crocodilians, and may represent an intriguing case of convergent evolution with mammals.

A Signaled Locomotor Avoidance Action Is Fully Represented in the Neural Activity of the Midbrain Tegmentum.

Animals, including humans, readily learn to avoid harmful and threatening situations by moving in response to cues that predict the threat (e.g., fire alarm, traffic light). During a negatively reinforced sensory-guided locomotor action, known as signaled active avoidance, animals learn to avoid a harmful unconditioned stimulus (US) by moving away when signaled by a harmless conditioned stimulus (CS) that predicts the threat. CaMKII-expressing neurons in the pedunculopontine tegmentum area (PPT) of the midbrain locomotor region have been shown to play a critical role in the expression of this learned behavior, but the activity of these neurons during learned behavior is unknown. Using calcium imaging fiber photometry in freely behaving mice, we show that PPT neurons sharply activate during presentation of the auditory CS that predicts the threat before onset of avoidance movement. PPT neurons activate further during the succeeding CS-driven avoidance movement, or during the faster US-driven escape movement. PPT neuron activation was weak during slow spontaneous movements but correlated sharply with movement speed and, therefore, with the urgency of the behavior. Moreover, using optogenetics, we found that these neurons must discharge during the signaled avoidance interval for naive mice to effectively learn the active avoidance behavior. As an essential hub for signaled active avoidance, neurons in the midbrain tegmentum process the conditioned cue that predicts the threat and discharge sharply relative to the speed or apparent urgency of the avoidance (learned) and escape (innate) responses.SIGNIFICANCE STATEMENT During signaled active avoidance behavior, subjects move away to avoid a threat when directed by an innocuous sensory stimulus. Using imaging methods in freely behaving mice, we found that the activity of neurons in a part of the midbrain, known as the pedunculopontime tegmentum, increases during the presentation of the innocuous sensory stimulus that predicts the threat and also during the expression of the learned behavior as mice move away to avoid the threat. In addition, inhibiting these neurons abolishes the ability of mice to learn the behavior. Thus, neurons in this part of the midbrain code and are essential for signaled active avoidance behavior.

Synaptic Adaptations at the Rostromedial Tegmental Nucleus Underlie Individual Differences in Cocaine Avoidance Behavior.

Although cocaine is powerfully rewarding, not all individuals are equally prone to abusing this drug. We postulate that these differences arise in part because some individuals exhibit stronger aversive responses to cocaine that protect them from cocaine seeking. Indeed, using conditioned place preference (CPP) and a runway operant cocaine self-administration task, we demonstrate that avoidance responses to cocaine vary greatly between individual high cocaine-avoider and low cocaine-avoider rats. These behavioral differences correlated with cocaine-induced activation of the rostromedial tegmental nucleus (RMTg), measured using both in vivo firing and c-fos, whereas slice electrophysiological recordings from ventral tegmental area (VTA)-projecting RMTg neurons showed that relative to low avoiders, high avoiders exhibited greater intrinsic excitability, greater transmission via calcium-permeable AMPA receptors (CP-AMPARs), and higher presynaptic glutamate release. In behaving animals, blocking CP-AMPARs in the RMTg with NASPM reduced cocaine avoidance. Hence, cocaine addiction vulnerability may be linked to multiple coordinated synaptic differences in VTA-projecting RMTg neurons.SIGNIFICANCE STATEMENT Although cocaine is highly addictive, not all individuals exposed to cocaine progress to chronic use for reasons that remain unclear. We find that cocaine's aversive effects, although less widely studied than its rewarding effects, show more individual variability, are predictive of subsequent propensity to seek cocaine, and are driven by variations in RMTg in response to cocaine that arise from distinct alterations in intrinsic excitability and glutamate transmission onto VTA-projecting RMTg neurons.

Clinical Correlation Between Vertical Gaze Palsy and Midbrain Volume in Progressive Supranuclear Palsy.

BACKGROUND: Supranuclear vertical gaze palsies and slowed vertical saccades are characteristic clinic features of progressive supranuclear palsy (PSP). The "hummingbird sign," reflective of midbrain atrophy, is a classic radiographic sign of PSP. Correlation between eye movement abnormalities and radiographic findings in PSP has been reported previously. However, due to the use of clinical criteria not commonly employed in neuro-ophthalmic practice and neuroimaging techniques that are not widely available, it remains unclear whether correlation between midbrain structure and characteristic ocular-motor disturbances can be helpful to neuro-ophthalmologists seeking to adjudicate difficult or unusual diagnostic cases. METHODS: Patients with a diagnosis of probable PSP according to Movement Disorders Society criteria were studied retrospectively. A neuroradiologist calculated brainstem volumes in enrolled participants and normal controls. Spearman correlations were used to correlate the extent of eye movement limitation as assessed by 2 neuro-ophthalmologists with brainstem volumes. RESULTS: Fourteen participants with PSP and 15 healthy controls with similar age and gender distribution were enrolled and evaluated retrospectively. All 14 participants with PSP had undergone MRIs. Midbrain atrophy significantly correlated with the PSP rating scale (P < 0.001). PSP patients had significantly reduced volumes in the midbrain (P -0.0026), tegmentum (0.0001), tectum (0.0001), and medulla (P = 0.0024) compared with normal controls. Notes documenting quantified ocular motor function were available in 7 of 14 participants with PSP. Midbrain atrophy significantly correlated with in the extent of upward gaze limitation (P = 0.03). CONCLUSIONS: The severity of upward gaze limitation correlates with the severity of midbrain atrophy in patients with PSP. Recognition of this correlation may help to adjudicate diagnostic dilemmas and guide further evaluation.

Categorical Signaling of the Strongest Stimulus by an Inhibitory Midbrain Nucleus.

The nucleus isthmi pars magnocellularis (Imc), a group of inhibitory neurons in the midbrain tegmentum, is a critical component of the spatial selection network in the vertebrate midbrain. It delivers long-range inhibition among different portions of the space map in the optic tectum (OT), thereby mediating stimulus competition in the OT. Here, we investigate the properties of relative strength-dependent competitive interactions within the Imc, in barn owls of both sexes. We find that when Imc neurons are presented simultaneously with one stimulus inside the receptive field and a second, competing stimulus outside, they exhibit gradual or switch-like response profiles as a function of relative stimulus strength. They do so both when the two stimuli are of the same sensory modality (both visual) or of different sensory modalities (visual and auditory). Moreover, Imc neurons signal the strongest stimulus in a dynamically flexible manner, indicating that Imc responses reflect an online comparison between the strengths of the competing stimuli. Notably, Imc neurons signal the strongest stimulus more categorically, and earlier than the OT. Paired recordings at spatially aligned sites in the Imc and OT reveal that although some properties of stimulus competition, such as the bias of competitive response profiles, are correlated, others such as the steepness of response profiles, are set independently. Our results demonstrate that the Imc is itself an active site of competition, and may be the first site in the midbrain selection network at which stimulus competition is resolved.SIGNIFICANCE STATEMENT This work sheds light on the functional properties of a small group of inhibitory neurons in the vertebrate midbrain that play a key part in how the brain selects a target among competitors. A better understanding of the functioning of these neurons is an important building block for the broader understanding of how distracters are suppressed, and of spatial attention and its dysfunction.

Distinct purinergic receptor-mediated currents of rat oculomotor integrator neurons characterized by different firing patterns.

The prepositus hypoglossi nucleus (PHN) and the interstitial nucleus of Cajal (INC) are oculomotor neural integrators involved in the control of horizontal and vertical gaze, respectively. We previously reported that local application of adenosine 5'-trisphosphate (ATP) to PHN neurons induced P2X receptor-mediated fast inward currents, P2Y receptor-mediated slow inward currents, and/or adenosine P1 receptor-mediated slow outward currents. In contrast to the findings on PHN neurons, the expression of functional purinergic receptors in INC neurons has not been examined. In this study, we investigated ATP-induced current responses in INC neurons and the distributions of the three current types across distinct firing patterns in PHN and INC neurons using whole cell recordings of rat brainstem slices. The application of ATP induced all three current types in INC neurons. Pharmacological analyses indicated that the fast inward and slow outward currents were mainly mediated by the P2X and P1 subtypes, respectively, corresponding to the receptor subtypes in PHN neurons. However, agonists of the P2Y subtype did not induce the slow inward current in INC neurons, suggesting that other subtypes or mechanisms are responsible for this current. Analysis of the distribution of the three current types in PHN and INC neurons revealed that the proportions of the currents were distinctly dependent on the firing patterns of PHN neurons whereas the proportion of the fast inward current was higher during all firing patterns of INC neurons. The different distributions of ATP-induced currents suggest distinct modes of purinergic modulation specific to horizontal and vertical integrators.NEW & NOTEWORTHY The roles of purinergic signaling on vertical (mediated by the interstitial nucleus of Cajal; INC) and horizontal (prepositus hypoglossal nucleus; PHN) gaze control are not understood. Here, we report three current types induced by ATP in INC neurons; the distribution of these current types across different types of INC neurons is different from that in PHN neurons. These results suggest distinct modes of purinergic modulation in horizontal and vertical gaze control centers.

Human wildtype tau expression in cholinergic pedunculopontine tegmental neurons is sufficient to produce PSP-like behavioural deficits and neuropathology.

Progressive Supranuclear Palsy (PSP) is the most common atypical parkinsonism and exhibits hallmark symptomology including motor function impairment and dysexecutive dementia. In contrast to Parkinson's disease, the underlying pathology displays aggregation of the protein tau, which is also seen in disorders such as Alzheimer's disease. Currently, there are no pharmacological treatments for PSP, and drug discovery efforts are hindered by the lack of an animal model specific to PSP. Based on previous results and clinical pathology, it was hypothesized that viral deposition of tau in cholinergic neurons within the hindbrain would produce a tauopathy along neural connections to produce PSP-like symptomology and pathology. By using a combination of ChAT-CRE rats and CRE-dependent AAV vectors, wildtype human tau (the PSP-relevant 1N4R isoform; hTau) was expressed in hindbrain cholinergic neurons. Compared to control subjects (GFP), rats with tau expression displayed deficits in a variety of behavioural paradigms: acoustic startle reflex, marble burying, horizontal ladder and hindlimb motor reflex. Postmortem, the hTau rats had significantly reduced number of cholinergic pedunculopontine tegmentum and dopaminergic substantia nigra neurons, as well as abnormal tau deposits. This preclinical model has multiple points of convergence with the clinical features of PSP, some of which distinguish between PSP and Parkinson's disease.

Laterodorsal tegmentum-ventral tegmental area projections encode positive reinforcement signals.

The laterodorsal tegmentum (LDT) is a brainstem nucleus classically involved in REM sleep and attention, and that has recently been associated with reward-related behaviors, as it controls the activity of ventral tegmental area (VTA) dopaminergic neurons, modulating dopamine release in the nucleus accumbens. To further understand the role of LDT-VTA inputs in reinforcement, we optogenetically manipulated these inputs during different behavioral paradigms in male rats. We found that in a two-choice instrumental task, optical activation of LDT-VTA projections shifts and amplifies preference to the laser-paired reward in comparison to an otherwise equal reward; the opposite was observed with inhibition experiments. In a progressive ratio task, LDT-VTA activation boosts motivation, that is, enhances the willingness to work to get the reward associated with LDT-VTA stimulation; and the reverse occurs when inhibiting these inputs. Animals abolished preference if the reward was omitted, suggesting that LDT-VTA stimulation adds/decreases value to the stimulation-paired reward. In addition, we show that LDT-VTA optical activation induces robust preference in the conditioned and real-time place preference tests, while optical inhibition induces aversion. The behavioral findings are supported by electrophysiological recordings and c-fos immunofluorescence correlates in downstream target regions. In LDT-VTA ChR2 animals, we observed an increase in the recruitment of lateral VTA dopamine neurons and D1 neurons from nucleus accumbens core and shell; whereas in LDT-VTA NpHR animals, D2 neurons appear to be preferentially recruited. Collectively, these data show that the LDT-VTA inputs encode positive reinforcement signals and are important for different dimensions of reward-related behaviors.

Repeated but Not Single Administration of Ketamine Prolongs Increases of the Firing Activity of Norepinephrine and Dopamine Neurons.

BACKGROUND: Clinical studies have shown that the rapid antidepressant effect of the glutamate N-methyl-D-aspartate receptor antagonist ketamine generally disappears within 1 week but can be maintained by repeated administration. Preclinical studies showed that a single ketamine injection immediately increases the firing and burst activity of norepinephrine (NE) neurons, but not that of serotonin (5-HT) neurons. It also enhances the population activity of dopamine (DA) neurons. In the present study, we investigated whether such alterations of monoamine neuronal firing are still present 1 day after a single injection, and whether they can be maintained by repeated injections. METHODS: Rats received a single ketamine injection or 6 over 2 weeks and the firing activity of dorsal raphe nucleus 5-HT, locus coeruleus NE, and ventral tegmental area DA neurons was assessed. RESULTS: One day following a single injection of ketamine, there was no change in the firing activity of 5-HT, NE, or DA neurons. One day after repeated ketamine administration, however, there was a robust increase of the firing activity of NE neurons and an enhancement of burst and population activities of DA neurons, but still no change in firing parameters of 5-HT neurons. The increased activity of NE neurons was no longer present 3 days after the last injection, whereas that of DA neurons was still present. DA neurons were firing normally 7 days after repeated injections. CONCLUSION: These results imply that the enhanced activity of NE and DA neurons may play a significant role in the maintenance of the antidepressant action of ketamine.

Cerebellar projections to the macaque midbrain tegmentum: Possible near response connections.

Since most gaze shifts are to targets that lie at a different distance from the viewer than the current target, gaze changes commonly require a change in the angle between the eyes. As part of this response, lens curvature must also be adjusted with respect to target distance by the ciliary muscle. It has been suggested that projections by the cerebellar fastigial and posterior interposed nuclei to the supraoculomotor area (SOA), which lies immediately dorsal to the oculomotor nucleus and contains near response neurons, support this behavior. However, the SOA also contains motoneurons that supply multiply innervated muscle fibers (MIFs) and the dendrites of levator palpebrae superioris motoneurons. To better determine the targets of the fastigial nucleus in the SOA, we placed an anterograde tracer into this cerebellar nucleus in Macaca fascicularis monkeys and a retrograde tracer into their contralateral medial rectus, superior rectus, and levator palpebrae muscles. We only observed close associations between anterogradely labeled boutons and the dendrites of medial rectus MIF and levator palpebrae motoneurons. However, relatively few of these associations were present, suggesting these are not the main cerebellar targets. In contrast, labeled boutons in SOA, and in the adjacent central mesencephalic reticular formation (cMRF), densely innervated a subpopulation of neurons. Based on their location, these cells may represent premotor near response neurons that supply medial rectus and preganglionic Edinger-Westphal motoneurons. We also identified lens accommodation-related cerebellar afferent neurons via retrograde trans-synaptic transport of the N2c rabies virus from the ciliary muscle. They were found bilaterally in the fastigial and posterior interposed nuclei, in a distribution which mirrored that of neurons retrogradely labeled from the SOA and cMRF. Our results suggest these cerebellar neurons coordinate elements of the near response during symmetric vergence and disjunctive saccades by targeting cMRF and SOA premotor neurons.

A nodal point for brain-state transitions: the mesopontine tegmental anesthesia area (MPTA) in mice.

The mesopontine tegmental anesthesia area (MPTA) was identified in rats as a singular brainstem locus at which microinjection of minute quantities of GABAergic agents rapidly and reversibly induces loss-of-consciousness and a state of general anesthesia, while lesioning renders animals insensitive to anesthetics at normal systemic doses. Obtaining similar results in mice has been challenging, however, slowing research progress on how anesthetics trigger brain-state transitions. We have identified roadblocks that impeded translation from rat to mouse and tentatively located the MPTA equivalent in this second species. We describe here a series of modifications to the rat protocol that allowed us to document pro-anesthetic changes in mice following localized stereotactic delivery of minute quantities (20 nL) of the GABAA-receptor agonist muscimol into the brainstem mesopontine tegmentum. The optimal locus identified proved to be homologous to the MPTA in rats, and local neuronal populations in rats and mice were similar in size and shape. This outcome should facilitate application of the many innovative gene-based methodologies available primarily in mice to the study of how activity in brainstem MPTA neurons brings about anesthetic loss-of-consciousness and permits pain-free surgery.

Acupuncture reduces cocaine psychomotor responses by activating the rostromedial tegmental nucleus.

The rostromedial tegmental nucleus (RMTg), a GABAergic afferent to midbrain dopamine (DA) neurons, has emerged as an integral player in both rewarding and nociceptive responses. While previous studies have demonstrated that acupuncture modulates DA transmission in the mesolimbic reward system originating in the ventral tegmental area (VTA) and projecting to the nucleus accumbens (NAc) and can reduce drug self-administration, the central links between peripheral acupuncture signals and brain reward systems are not well-characterized. Thus, we hypothesised that acupuncture would elicit inhibitory signals from RMTg neurons to brain reward systems. Acupuncture reduced acute cocaine-induced locomotor activity and DA release in a point-specific manner, which was blocked by optogenetic silencing or chemical lesion of the RMTg. The acupuncture effect was mimicked by chemical activation of the RMTg. Acupuncture activated RMTg GABA neurons. In addition, the inhibitory effects of acupuncture on acute cocaine-induced locomotor activity were prevented by electrolytic lesions of the lateral habenula (LHb) or fasciculus retroflexus (FR), areas known to project to the RMTg. These findings suggest that acupuncture recruits the RMTg to reduce the psychomotor responses enhanced by acute cocaine.

A neural pathway controlling motivation to exert effort.

The neural mechanisms conferring reduced motivation, as observed in depressed individuals, is poorly understood. Here, we examine in rodents if reduced motivation to exert effort is controlled by transmission from the lateral habenula (LHb), a nucleus overactive in depressed-like states, to the rostromedial tegmental nucleus (RMTg), a nucleus that inhibits dopaminergic neurons. In an aversive test wherein immobility indicates loss of effort, LHb→RMTg transmission increased during transitions into immobility, driving LHb→RMTg increased immobility, and inhibiting LHb→RMTg produced the opposite effects. In an appetitive test, driving LHb→RMTg reduced the effort exerted to receive a reward, without affecting the reward's hedonic property. Notably, LHb→RMTg stimulation only affected specific aspects of these motor tasks, did not affect all motor tasks, and promoted avoidance, indicating that LHb→RMTg activity does not generally reduce movement but appears to carry a negative valence that reduces effort. These results indicate that LHb→RMTg activity controls the motivation to exert effort and may contribute to the reduced motivation in depression.

A Confocal Microscopic Study of Gene Transfer into the Mesencephalic Tegmentum of Juvenile Chum Salmon, Oncorhynchus keta, Using Mouse Adeno-Associated Viral Vectors.

To date, data on the presence of adenoviral receptors in fish are very limited. In the present work, we used mouse recombinant adeno-associated viral vectors (rAAV) with a calcium indicator of the latest generation GCaMP6m that are usually applied for the dorsal hippocampus of mice but were not previously used for gene delivery into fish brain. The aim of our work was to study the feasibility of transduction of rAAV in the mouse hippocampus into brain cells of juvenile chum salmon and subsequent determination of the phenotype of rAAV-labeled cells by confocal laser scanning microscopy (CLSM). Delivery of the gene in vivo was carried out by intracranial injection of a GCaMP6m-GFP-containing vector directly into the mesencephalic tegmentum region of juvenile (one-year-old) chum salmon, Oncorhynchus keta. AAV incorporation into brain cells of the juvenile chum salmon was assessed at 1 week after a single injection of the vector. AAV expression in various areas of the thalamus, pretectum, posterior-tuberal region, postcommissural region, medial and lateral regions of the tegmentum, and mesencephalic reticular formation of juvenile O. keta was evaluated using CLSM followed by immunohistochemical analysis of the localization of the neuron-specific calcium binding protein HuCD in combination with nuclear staining with DAPI. The results of the analysis showed partial colocalization of cells expressing GCaMP6m-GFP with red fluorescent HuCD protein. Thus, cells of the thalamus, posterior tuberal region, mesencephalic tegmentum, cells of the accessory visual system, mesencephalic reticular formation, hypothalamus, and postcommissural region of the mesencephalon of juvenile chum salmon expressing GCaMP6m-GFP were attributed to the neuron-specific line of chum salmon brain cells, which indicates the ability of hippocampal mammal rAAV to integrate into neurons of the central nervous system of fish with subsequent expression of viral proteins, which obviously indicates the neuronal expression of a mammalian adenoviral receptor homolog by juvenile chum salmon neurons.

Circuit-Specific Early Impairment of Proprioceptive Sensory Neurons in the SOD1G93A Mouse Model for ALS.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease in which motor neurons degenerate, resulting in muscle atrophy, paralysis, and fatality. Studies using mouse models of ALS indicate a protracted period of disease development with progressive motor neuron pathology, evident as early as embryonic and postnatal stages. Key missing information includes concomitant alterations in the sensorimotor circuit essential for normal development and function of the neuromuscular system. Leveraging unique brainstem circuitry, we show in vitro evidence for reflex circuit-specific postnatal abnormalities in the jaw proprioceptive sensory neurons in the well-studied SOD1G93A mouse. These include impaired and arrhythmic action potential burst discharge associated with a deficit in Nav1.6 Na+ channels. However, the mechanoreceptive and nociceptive trigeminal ganglion neurons and the visual sensory retinal ganglion neurons were resistant to excitability changes in age-matched SOD1G93A mice. Computational modeling of the observed disruption in sensory patterns predicted asynchronous self-sustained motor neuron discharge suggestive of imminent reflexive defects, such as muscle fasciculations in ALS. These results demonstrate a novel reflex circuit-specific proprioceptive sensory abnormality in ALS.SIGNIFICANCE STATEMENT Neurodegenerative diseases have prolonged periods of disease development and progression. Identifying early markers of vulnerability can therefore help devise better diagnostic and treatment strategies. In this study, we examined postnatal abnormalities in the electrical excitability of muscle spindle afferent proprioceptive neurons in the well-studied SOD1G93A mouse model for neurodegenerative motor neuron disease, amyotrophic lateral sclerosis. Our findings suggest that these proprioceptive sensory neurons are exclusively afflicted early in the disease process relative to sensory neurons of other modalities. Moreover, they presented Nav1.6 Na+ channel deficiency, which contributed to arrhythmic burst discharge. Such sensory arrhythmia could initiate reflexive defects, such as muscle fasciculations in amyotrophic lateral sclerosis, as suggested by our computational model.

Presynaptic dopamine deficit in minimally conscious state patients following traumatic brain injury.

Dopaminergic stimulation has been proposed as a treatment strategy for post-traumatic brain injured patients in minimally conscious state based on a clinical trial using amantadine, a weak dopamine transporter blocker. However, a specific contribution of dopaminergic neuromodulation in minimally conscious state is undemonstrated. In a phase 0 clinical trial, we evaluated 13 normal volunteers and seven post-traumatic minimally conscious state patients using 11C-raclopride PET to estimate dopamine 2-like receptors occupancy in the striatum and central thalamus before and after dopamine transporter blockade with dextroamphetamine. If a presynaptic deficit was observed, a third and a fourth 11C-raclopride PET were acquired to evaluate changes in dopamine release induced by l-DOPA and l-DOPA+dextroamphetamine. Permutation analysis showed a significant reduction of dopamine release in patients, demonstrating a presynaptic deficit in the striatum and central thalamus that could not be reversed by blocking the dopamine transporter. However, administration of the dopamine precursor l-DOPA reversed the presynaptic deficit by restoring the biosynthesis of dopamine from both ventral tegmentum and substantia nigra. The advantages of alternative pharmacodynamic approaches in post-traumatic minimally conscious state patients should be tested in clinical trials, as patients currently refractory to amantadine might benefit from them.

Brain region-specific neuromedin U signalling regulates alcohol-related behaviours and food intake in rodents.

Albeit neuromedin U (NMU) attenuates alcohol-mediated behaviours, its mechanisms of action are poorly defined. Providing that the behavioural effects of alcohol are processed within the nucleus accumbens (NAc) shell, anterior ventral tegmental area (aVTA), and laterodorsal tegmental area (LDTg), we assessed the involvement of NMU signalling in the aforementioned areas on alcohol-mediated behaviours in rodents. We further examined the expression of NMU and NMU receptor 2 (NMUR2) in NAc and the dorsal striatum of high compared with low alcohol-consuming rats, as this area is of importance in the maintenance of alcohol use disorder (AUD). Finally, we investigated the involvement of NAc shell, aVTA and LDTg in the consumption of chow and palatable peanut butter, to expand the link between NMU and reward-related behaviours. We demonstrated here, that NMU into the NAc shell, but not aVTA or LDTg, blocked the ability of acute alcohol to cause locomotor stimulation and to induce memory retrieval of alcohol reward, as well as reduced peanut butter in mice. In addition, NMU into NAc shell decreased alcohol intake in rats. On a molecular level, we found increased NMU and decreased NMUR2 expression in the dorsal striatum in high compared with low alcohol-consuming rats. Both aVTA and LDTg, rather than NAc shell, were identified as novel sites of action for NMU's anorexigenic properties in mice based on NMU's ability to selectively reduce chow intake when injected to these areas. Collectively, these data indicate that NMU signalling in different brain areas selectively modulates different behaviours.