Biology subjects

Chee, M. J.

Publications and source records attributed to Chee, M. J..

9 recordsLinked to original sources

bioRxiv2026

Intersectional genetic targeting of molecularly distinct melanin-concentrating hormone neuron subpopulations reveals distinct and overlapping subcircuit architecture

The lateral hypothalamic area (LHA) coordinates crucial and innate behaviors through heterogeneous yet poorly understood neuronal populations and circuits. Neurons that produce melanin-concentrating hormone (MCH; encoded by the Pmch gene) are primarily localized in the LHA and contribute to feeding, sleep-wake regulation, memory, and reward-associated behaviors. Building on foundational anatomical studies, our single-cell transcriptomic analysis of the LHA identified markers distinguishing at least two transcriptionally distinct MCH subpopulations. These subpopulations are distinguished by the selective enrichment of the Tacr3 and Cartpt transcripts, with each comprising approximately half of the total MCH population. However, these subpopulations remain poorly understood owing to limited genetic targeting tools. We hypothesized that MCH neurons operate through functionally distinct, parallel subcircuits that exhibit unique neuroanatomical projections. In this study, we leveraged Pmch and Tacr3 expression to intersectionally dissociate these neurons into Type I MCH (Pmch+/Tacr3+) and Type II MCH (Pmch+/Tacr3-) subpopulations. We first validated a Tacr3-Flpo mouse line for the genetic targeting of the Type I MCH subpopulation. We then generated an intersectional Type I MCH neuron reporter line to visualize Type I axonal projections. In parallel, we injected a dual recombinase mouse line with the intersectional genetic targeting viruses, Conditional Viral Expression by Ribozyme Guided Degradation (ConVERGD), and identified overlapping and distinct projections by Type I and Type II MCH neurons using fluorescence microscopy, super-resolution imaging, and high-spatial resolution fiber mapping. Overall, these data establish the first subpopulation-resolved anatomical framework for molecularly distinct Type I and Type II MCH subcircuits, thus establishing a foundation for future circuit-level functional studies.

neuroscience↗

bioRxiv2024

Anorectic and anxiogenic actions of cocaine- and amphetamine-regulated transcript in the lateral septum

Cocaine-and amphetamine-regulated transcript (CART) is produced in several brain regions including the hypothalamus where it is made in cells that also produce melanin-concentrating hormone (MCH). MCH+CART cells densely innervate the lateral septum (LS), which integrates food- and mood-related behaviours. However, while MCH typically promotes feeding and anxiolysis, CART suppresses feeding and promotes anxiogenesis. The LS is a target site of the orexigenic actions of MCH, but it is not known if the actions of CART converge or oppose that of MCH in the LS. We implanted a bilateral cannula over the lateral or central LS of male and female wildtype mice and infused vehicle, CART55-102, or co-infused CART and MCH. CART did not alter chow intake but suppressed the intake of a palatable high sugar diet in male and female mice, especially when delivered in the medial LS. Furthermore, CART also prevented the orexigenic effect of MCH on palatable feeding intake when infused in the medial LS. We then assessed if CART regulated anxiety-like behaviour via the LS and found that intra-LS CART infusion reduced time spent in the center of an open field in male but not female mice. Our findings indicated that CART elicited anorectic and anxiogenic actions and may function in opposition to or independently of MCH in the LS. These outcomes suggested that putative CART and MCH co-release from MCH neurons may provide biphasic regulation of feeding and anxiety via the LS.

neuroscience↗

bioRxiv2024

Brain-wide projections of mouse dopaminergic zona incerta neurons

The zona incerta (ZI) supports diverse behaviors including binge feeding, sleep/wake cycles, nociception, and hunting. This diversity of functions can be attributed to the heterogenous neurochemicals, cytoarchitecture, and efferent connections that characterize the ZI. The ZI is predominantly GABAergic, but we recently identified a subset of medial ZI GABA cells that co-express dopamine (DA), as marked by the enzyme tyrosine hydroxylase (TH). While the role of GABA within the ZI is well studied, little is understood about the function of ZI DA cells. To identify potential roles of ZI DA cells we mapped the efferent fiber projections from Th-cre ZI cells. We first validated a Th-cre;L10-Egfp mouse line and found that medial Egfp ZI cells were more likely to co-express TH-immunoreactivity (TH-ir). We thus delivered a cre-dependent virus into the medial ZI of Th-cre or Th-cre;L10-Egfp mice and selected two injection cases for full brain mapping. We selected the cases with the lowest (17%) and highest (53%) percentage of colocalization between TH-ir and virus transfected cells labelled with DsRed. Overall, DsRed-labelled fibers were observed throughout the brain and were most prominent within motor-related regions of the midbrain (MBmot), notably the periaqueductal grey area and superior colliculus. We also observed considerable DsRed-labelled fibers within the polymodal cortex associated regions of the thalamus (DORpm), including the paraventricular thalamic nucleus and nucleus of reunions. Overall, ZI DA cells displayed a similar connectivity profile to ZI GABA cells, suggesting that ZI DA cells may perform synergistic or opposing functions at the same target sites. List of RRIDsAB_2201528, AB_10013483, AB_11177031, AB_2340593, AB_2315778, SCR_016477 Three key points- Tyrosine hydroxylase immunoreactivity was more prominent in the medial zona incerta - Intersection points from two Th-cre injection cases revealed common target regions - Dopaminergic zona incerta cells project predominantly to motor-related brain regions

neuroscience↗

bioRxiv2024

Melanin-concentrating hormone promotes feeding through the lateral septum

Feeding is necessary for survival but can be hindered by anxiety or fear, thus neural systems that can regulate anxiety states are key to elucidating the expression of food-related behaviors. Melanin-concentrating hormone (MCH) is a neuropeptide produced in the lateral hypothalamus that promotes feeding and anxiogenesis. The orexigenic actions of MCH that prolong ongoing homeostatic or hedonic feeding are context-dependent and more prominent in male than female rodents, but it is not clear where MCH acts to initiate feeding. The lateral septum (LS) promotes feeding and suppresses anxiogenesis when inhibited, and it comprises the densest projections from MCH neurons. However, it is not known whether the LS is a major contributor to MCH-mediated feeding. As MCH inhibits LS cells by MCH receptor (MCHR1) activation, MCH may promote feeding via the LS. We bilaterally infused MCH into the LS and found that MCH elicited a rapid and long-lasting increase in the consumption of standard chow and a palatable, high sugar diet in male and female mice; these MCH effects were blocked by the co-administration of a MCHR1 antagonist TC-MCH 7c. Interestingly, the orexigenic effect of MCH was abolished in a novel, anxiogenic environment even when presented with a food reward, but MCH did not induce anxiety-like behaviors. These findings indicated the LS as a novel region underlying orexigenic MCH actions, which stimulated and enhanced feeding in both sexes in a context -dependent manner that was most prominent in the homecage.

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bioRxiv2023

Inhibitory actions of melanin-concentrating hormone in the lateral septum

Melanin-concentrating hormone (MCH) neurons can coexpress several neuropeptides or neurotransmitters and send widespread projections throughout the brain. Notably, there is a dense cluster of nerve terminals from MCH neurons in the lateral septum (LS) that innervate LS cells by glutamate release. The LS is also a key region integrating stress- and anxiety-like behaviours that are also emerging roles of MCH neurons. However, it is not known if the MCH peptide acts within the LS or whether MCH target sites are localized. We analysed the projections from MCH neurons in male and female mice anteroposteriorly throughout the LS and found spatial overlap between the distribution pattern of MCH-immunoreactive (MCH-ir) fibers with MCH receptor Mchr1 mRNA hybridization or MCHR1-ir cells. This overlap was most prominent along the ventral and lateral border of the rostral part of the LS (LSr). Most MCHR1-labeled LS neurons laid adjacent to passing MCH-ir fibers, but some MCH-ir varicosities directly contacted the soma or cilium of MCHR1-labeled LS neurons. We thus performed whole-cell patch-clamp recordings from MCHR1-rich LSr regions to determine if and how LS cells respond to MCH. Bath application of MCH to acute brain slices activated a bicuculline-sensitive chloride current that directly hyperpolarized LS cells. This MCH-mediated hyperpolarization was blocked by calphostin C and suggested that the inhibitory actions of MCH were mediated by protein kinase C-dependent activation of GABAA receptors. Taken together, these findings defined potential hotspots within the LS that may elucidate the contributions of MCH to stress- or anxiety-related feeding behaviours. Key pointsO_LIRESEARCH QUESTION. Melanin-concentrating hormone (MCH) neurons have dense nerve terminals within the lateral septum (LS), a key region underlying stress- and anxiety-like behaviours that are emerging roles of the MCH system, but it is not known if the LS is a MCH target site. C_LIO_LINEUROANATOMY. We found spatial overlap between MCH-immunoreactive fibers, Mchr1 mRNA, and MCHR1 protein expression especially along the lateral border of the LS. C_LIO_LIELECTROPHYSIOLOGY. Within MCHR1-rich regions, MCH directly inhibited LS cells by increasing a chloride conductance in a protein kinase C-dependent manner. C_LIO_LISIGNIFICANCE. Electrophysiological MCH effects in brain slices have been elusive and even fewer have described the mechanisms of MCH action. Our findings demonstrated, to our knowledge, the first description of MCHR1 Gq-coupling in brain slices, which was previously predicted in cell or primary culture models only. Together, these findings defined hotspots and mechanistic underpinnings for MCH effects such as in stress- and anxiety-related behaviours. C_LI

neuroscience↗

bioRxiv2023

Neuroanatomical, electrophysiological, and morphological characterization of melanin-concentrating hormone cells coexpressing cocaine- and amphetamine-regulated transcript

Melanin-concentrating hormone (MCH) cells in the hypothalamus regulate fundamental physiological functions like energy balance, sleep, and reproduction. This diversity may be ascribed to the neurochemical heterogeneity among MCH cells. One prominent subpopulation of MCH cells coexpresses cocaine- and amphetamine-regulated transcript (CART), and as MCH and CART can have opposing actions, MCH/CART+ and MCH/CART- cells may differentially modulate behavioural outcomes. However, it is not known if there are differences in cellular properties underlying their functional differences, thus we compared the neuroanatomical, electrophysiological, and morphological properties of MCH cells in male and female Mch-cre;L10-Egfp reporter mice. Half of MCH cells expressed CART and were most prominent in the medial hypothalamus. Whole-cell patch-clamp recordings revealed differences in their passive and active membrane properties in a sex-dependent manner. Female MCH/CART+ cells had lower input resistances, but male cells largely differed in their firing properties. All MCH cells increased firing when stimulated, but their firing frequency decreases with sustained stimulation. MCH/CART+ cells showed stronger spike rate adaptation than MCH/CART- cells. The kinetics of excitatory events at MCH cells also differed by cell type, as the rising rate of excitatory events was slower at MCH/CART+ cells. By reconstructing the dendritic arborization of our recorded cells, we found no sex differences, but male MCH/CART+ cells had less dendritic length and fewer branch points. Overall, distinctions in topographical division and cellular properties between MCH cells add to their heterogeneity and help elucidate their response to stimuli or effect on modulating their respective neural networks.

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bioRxiv2023

Regulation of the mouse ventral tegmental area by melanin-concentrating hormone

Melanin-concentrating hormone (MCH) acts via its sole receptor MCHR1 in rodents and is an important regulator of homeostatic behaviors like feeding, sleep, and mood to impact overall energy balance. The loss of MCH signaling by MCH or MCHR1 deletion produces hyperactive mice with increased energy expenditure, and these effects are consistently associated with a hyperdopaminergic state. We recently showed that MCH suppresses dopamine release in the nucleus accumbens, which principally receives dopaminergic projections from the ventral tegmental area (VTA), but the mechanisms underlying MCH-regulated dopamine release are not clearly defined. MCHR1 expression is widespread and includes dopaminergic VTA cells. However, as the VTA is a neurochemically diverse structure, we assessed Mchr1 gene expression at glutamatergic, GABAergic, and dopaminergic VTA cells and determined if MCH inhibited the activity of VTA cells and/or their local microcircuit. Mchr1 expression was robust in major VTA cell types, including most dopaminergic (78%) or glutamatergic cells (52%) and some GABAergic cells (38%). Interestingly, MCH directly inhibited dopaminergic and GABAergic cells but did not regulate the activity of glutamatergic cells. Rather, MCH produced a delayed increase in excitatory input to dopamine cells and a corresponding decrease in GABAergic input to glutamatergic VTA cells. Our findings suggested that MCH may acutely suppress dopamine release while disinhibiting local glutamatergic signaling to restore dopamine levels. This indicated that the VTA is a target of MCH action, which may provide bidirectional regulation of energy balance. Significance StatementRole of melanin-concentrating hormone (MCH) on energy balance may converge on the dopamine system via the mesolimbic pathway, as loss of MCH or MCH receptor (MCHR1) signaling increases hyperactivity and energy expenditure associated with a hyperdopaminergic state. MCH can suppress dopamine release within the mesolimbic pathway, but its underlying mechanism is not known. We thus determined if MCH could inhibit dopamine release through direct actions within the ventral tegmental area (VTA). We found that MCH directly inhibited dopaminergic VTA cells, but MCH also disinhibited excitatory input to dopamine cells. Therefore, we showed that the VTA is a putative target site supporting dopamine-dependent actions of MCH.

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bioRxiv2019

The distribution of neuronal primary cilia immunoreactive to melanin-concentrating hormone receptor 1 (MCHR1) in the murine prosencephalon

Melanin-concentrating hormone (MCH) is a ubiquitous vertebrate neuropeptide predominantly synthesized by neurons of the diencephalon that can act through two G protein-coupled receptors, called MCHR1 and MCHR2. The expression of Mchr1 has been investigated in both rats and mice, but its synthesis remains poorly described. After identifying an antibody that detects MCHR1 with high specificity, we employed immunohistochemistry to map the distribution of MCHR1 in the CNS of rats and mice. Multiple neurochemical markers were also employed to characterize some of the neuronal populations that synthesize MCHR1. Our results show that MCHR1 is abundantly found in a sensory subcellular structure called the neuronal primary cilium, which has been associated with the detection of free neurochemical agents released to act through volume transmission. Ciliary MCHR1 was found in a wide range of areas, including the olfactory bulb, cortical mantle, striatum, hippocampal formation, amygdala, midline thalamic nuclei, periventricular hypothalamic nuclei, and midbrain areas. No differences were observed between male and female mice, and rats and mice diverged in two key areas: the caudate-putamen nucleus and the subgranular zone of the dentate gyrus. Ciliary MCHR1 was found in close association to several neurochemical markers, including tyrosine hydroxylase, calretinin, kisspeptin, estrogen receptor, oxytocin, vasopressin, and corticotropin-releasing factor. Given the role of neuronal primary cilia in sensing free neurochemical messengers in the extracellular fluid, the widespread distribution of ciliary MCHR1, and the diverse neurochemical populations who synthesize MCHR1, our data indicates that volume transmission may play a prominent role in the normal function of the MCH system.

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bioRxiv2019

Distributions of hypothalamic neuron populations co-expressing 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 glutamate or GABA. We used Cre/loxP recombination to express enhanced GFP fluorescence (EGFP) in neurons that express the vesicular glutamate (vGLUT2) or GABA transporter (vGAT), then determined TH immunoreactivity in glutamatergic or GABAergic neurons, respectively. EGFP-positive vGLUT2 neurons were not TH-ir. However, discrete TH-ir signals colocalized with EGFP-positive vGAT neurons, which we validated by in situ hybridization for Vgat mRNA. In order to contextualize the observed pattern of TH+EGFP colocalization in vGAT neurons, we first performed Nissl-based parcellation and plane-of-section analysis, and then mapped the distribution of TH-ir vGAT neurons onto atlas templates from the Allen Reference Atlas (ARA) of the mouse brain. TH-ir vGAT 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 very strong presence of EGFP fluorescence in TH-ir neurons across all brain regions, but the most striking colocalization was found in the zona incerta (ZI) - a region assigned to the hypothalamus in the ARA - where every TH-ir neuron was EGFP-positive. Neurochemical characterization of these ZI neurons revealed that they display immunoreactivity for dopamine but not dopamine {beta}-hydroxylase. In aggregate, these findings indicate the existence of a novel hypothalamic population in the mouse that may signal through the release of GABA and/or dopamine.

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