Philosophical transactions of the Royal Society of London. Series B, Biological sciences
Frehner, SS;Dooley, KT;Palumbo, MC;Smith, AL;Goodman, MM;Bales, KL;Freeman, SM;
PMID: 35858098 | DOI: 10.1098/rstb.2021.0118
Oxytocin is an endogenous neuropeptide hormone that influences social behaviour and bonding in mammals. Variations in oxytocin receptor (OXTR) expression may play a role in the social deficits seen in autism spectrum disorder. Previous studies from our laboratory found a dense population of OXTR in the human substantia nigra (SN), a basal ganglia structure in the midbrain that is important in both movement and reward pathways. Here, we explore whether differences in OXTR can be identified in the dopaminergic SN pars compacta of individuals with autism. Postmortem human brain tissue specimens were processed for OXTR autoradiography from four groups: males with autism, females with autism, typically developing (TD) males and TD females. We found that females with autism had significantly lower levels of OXTR than the other groups. To examine potential gene expression differences, we performed in situ hybridization in adjacent slides to visualize and quantify OXTR mRNA as well as mRNA for tyrosine hydroxylase. We found no differences in mRNA levels for either gene across the four groups. These results suggest that a dysregulation in local OXTR protein translation or increased OXTR internalization/recycling may contribute to the differences in social symptoms seen in females with autism. This article is part of the theme issue 'Interplays between oxytocin and other neuromodulators in shaping complex social behaviours'.
Skiteva, O;Yao, N;Mantas, I;Zhang, X;Perlmann, T;Svenningsson, P;Chergui, K;
PMID: 37029193 | DOI: 10.1038/s41531-023-00500-5
In Parkinson's disease (PD), axons of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNc) degenerate before their cell bodies. Calcium influx during pacemaker firing might contribute to neuronal loss, but it is not known if dysfunctions of voltage-gated calcium channels (VGCCs) occur in DA neurons somata and axon terminals. We investigated T-type and L-type VGCCs in SNc-DA neurons of two mouse models of PD: mice with a deletion of the Nurr1 gene in DA neurons from an adult age (cNurr1 mice), and mice bearing the G2019S mutation in the gene coding for LRRK2 (G2019S mice). Adult cNurr1 mice displayed motor and DA deficits, while middle-aged G2019S mice did not. The number and morphology of SNc-DA neurons, most of their intrinsic membrane properties and pacemaker firing were unaltered in cNurr1 and G2019S mice compared to their control and wild-type littermates. L-type VGCCs contributed to the pacemaker firing of SNc-DA neurons in G2019S mice, but not in control, wild-type, and cNurr1 mice. In cNurr1 mice, but not G2019S mice, the contribution of T-type VGCCs to the pacemaker firing of SNc-DA neurons was reduced, and somatic dopamine-D2 autoreceptors desensitized more. Altered contribution of L-type and T-type VGCCs to the pacemaker firing was not observed in the presence of a LRRK2 kinase inhibitor in G2019S mice, and in the presence of a flavonoid with antioxidant activity in G2019S and cNurr1 mice. The role of L-type and T-type VGCCs in controlling dopamine release from axon terminals in the striatum was unaltered in cNurr1 and G2019S mice. Our findings uncover opposite changes, linked to oxidative stress, in the function of two VGCCs in DA neurons somata, but not axon terminals, in two different experimental PD models.
Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism
Mesa-Ciller, C;Turiel, G;Guajardo-Grence, A;Lopez-Rodriguez, AB;Egea, J;De Bock, K;Aragonés, J;Urrutia, AA;
PMID: 35929074 | DOI: 10.1177/0271678X221118236
A central response to insufficient cerebral oxygen delivery is a profound reprograming of metabolism, which is mainly regulated by the Hypoxia Inducible Factor (HIF). Among other responses, HIF induces the expression of the atypical mitochondrial subunit NDUFA4L2. Surprisingly, NDUFA4L2 is constitutively expressed in the brain in non-hypoxic conditions. Analysis of publicly available single cell transcriptomic (scRNA-seq) data sets coupled with high-resolution multiplexed fluorescence RNA in situ hybridization (RNA F.I.S.H.) revealed that in the murine and human brain NDUFA4L2 is exclusively expressed in mural cells with the highest levels found in pericytes and declining along the arteriole-arterial smooth muscle cell axis. This pattern was mirrored by COX4I2, another atypical mitochondrial subunit. High NDUFA4L2 expression was also observed in human brain pericytes in vitro, decreasing when pericytes are muscularized and further induced by HIF stabilization in a PHD2/PHD3 dependent manner. In vivo, Vhl conditional inactivation in pericyte targeting Ng2-cre transgenic mice dramatically induced NDUFA4L2 expression. Finally NDUFA4L2 inactivation in pericytes increased oxygen consumption and therefore the degree of HIF pathway induction in hypoxia. In conclusion our work reveals that NDUFA4L2 together with COX4I2 is a key hypoxic-induced metabolic marker constitutively expressed in pericytes coupling mitochondrial oxygen consumption and cellular hypoxia response.
Cline, MM;Juarez, B;Hunker, A;Regiarto, EG;Hariadi, B;Soden, ME;Zweifel, LS;
PMID: 36927614 | DOI: 10.7554/eLife.83760
The axonal guidance cue netrin-1 serves a critical role in neural circuit development by promoting growth cone motility, axonal branching, and synaptogenesis. Within the adult mouse brain, expression of the gene encoding (Ntn1) is highly enriched in the ventral midbrain where it is expressed in both GABAergic and dopaminergic neurons, but its function in these cell types in the adult system remains largely unknown. To address this, we performed viral-mediated, cell-type specific CRISPR-Cas9 mutagenesis of Ntn1 in the ventral tegmental area (VTA) of adult mice. Ntn1 loss-of-function in either cell type resulted in a significant reduction in excitatory postsynaptic connectivity. In dopamine neurons, the reduced excitatory tone had a minimal phenotypic behavioral outcome; however, reduced glutamatergic tone on VTA GABA neurons induced behaviors associated with a hyperdopaminergic phenotype. Simultaneous loss of Ntn1 function in both cell types largely rescued the phenotype observed in the GABA-only mutagenesis. These findings demonstrate an important role for Ntn1 in maintaining excitatory connectivity in the adult midbrain and that a balance in this connectivity within two of the major cell types of the VTA is critical for the proper functioning of the mesolimbic system.
Connexin mRNA distribution in adult mouse kidneys
Pflugers Archiv : European journal of physiology
Geis, L;Boudriot, FF;Wagner, C;
PMID: 34365513 | DOI: 10.1007/s00424-021-02608-0
Kidneys are thought to express eight different connexin isoforms (i.e., Cx 26, 30, 32, 37, 40, 43, 45, and 46), which form either hemichannels or gap junctions serving to intercellular communication and functional synchronization. Proper function of connexins has already been shown to be crucial for regulation of renal hemodynamics and renin secretion, and there is also growing evidence for connexins to play a role in pathologic conditions such as renal fibrosis or diabetic nephropathy. Therefore, exact intrarenal localization of the different connexin isoforms gains particular interest. Until now intrarenal expression of connexins has mainly been examined by immunohistochemistry, which in part generated conflicting results depending on antibodies and fixation protocols used. In this work, we used fluorescent RNAscope as an alternative technical approach to localize renal connexin mRNAs in healthy mouse kidneys. Addition of RNAscope probes for cell type specific mRNAs was used to assign connexin mRNA signals to specific cell types. We hereby found Cx26 mRNA strongly expressed in proximal tubules, Cx30 mRNA was selectively detected in the urothelium, and Cx32 mRNA was found in proximal tubules and to a lesser extent also in collecting ducts. Cx37 mRNA was mainly associated with vascular endothelium, Cx40 mRNA was largely found in glomerular mesangial and less in vascular endothelial cells, Cx43 mRNA was sparsely expressed by interstitial cells of all kidney zones, and Cx45 mRNA was predominantly found in smooth muscle cell layers of both blood vessels and ureter as well as in mesangial and interstitial (fibroblastic) cells. Cx46 mRNA could not be detected. In summary our results essentially confirm previous data on connexin expression in the renal vasculature and in glomeruli. In addition, they demonstrate strong connexin gene expression in proximal tubules, and they suggest significant connexin expression in resident tubulointerstitial cells.
Godefroy, D;Boukhzar, L;Mallouki, BY;Carpentier, E;Dubessy, C;Chigr, F;Tillet, Y;Anouar, Y;
PMID: 35066506 | DOI: 10.1159/000522091
Selenoprotein T (SELENOT), a PACAP-regulated thioredoxin-like protein, plays a role in catecholamine secretion and protects dopaminergic neurons. However, the role of SELENOT in the establishment of the catecholaminergic (CA) neuronal system is not known yet.We analyzed by immunohistochemistry and RNAscope in situ hybridization the distribution of SELENOT and the expression of its mRNA, respectively. In addition, 3D imaging involving immunostaining in toto, clearing through the iDISCO+ method, acquisitions by light sheet microscopy and processing of 3D images was performed to map the CA neuronal system. A semi-automatic quantification of 3D images was carried out.SELENOT protein and mRNA are widely distributed in the mouse brain, with important local variations. Three-dimensional mapping, through tyrosine hydroxylase (TH) labeling, and semi-automated quantification of CA neurons in brain-specific SELENOT knockout mice showed a significant decrease in the number of TH-positive neurons in the area postrema (AP-A2), the A11 cell group (A11) and the zona incerta (ZI-A13) of SELENOT-deficient females, and in the hypothalamus (Hyp-A12-A14-A15) of SELENOT-deficient females and males.These results showed that SELENOT is diffusely expressed in the mouse brain and that its deficiency impacts CA neuron distribution in different brain areas including Hyp-A12-A14-A15, in both male and female mice.S. Karger AG, Basel.
Pflugers Archiv : European journal of physiology
Heinl, ES;Broeker, KA;Lehrmann, C;Heydn, R;Krieger, K;Ortmaier, K;Tauber, P;Schweda, F;
PMID: 36480070 | DOI: 10.1007/s00424-022-02774-9
The natriuretic peptides (NPs) ANP (atrial natriuretic peptide) and BNP (B-type natriuretic peptide) mediate their widespread effects by activating the natriuretic peptide receptor-A (NPR-A), while C-type natriuretic peptide (CNP) acts via natriuretic peptide receptor-B (NPR-B). NPs are removed from the circulation by internalization via the natriuretic peptide clearance receptor natriuretic peptide receptor-C (NPR-C). In addition to their well-known functions, for instance on blood pressure, all three NPs confer significant cardioprotection and renoprotection. Since neither the NP-mediated renal functions nor the renal target cells of renoprotection are completely understood, we performed systematic localization studies of NP receptors using in situ hybridization (RNAscope) in mouse kidneys. NPR-A mRNA is highly expressed in glomeruli (mainly podocytes), renal arterioles, endothelial cells of peritubular capillaries, and PDGFR-receptor β positive (PDGFR-β) interstitial cells. No NPR-A mRNA was detected by RNAscope in the tubular system. In contrast, NPR-B expression is highest in proximal tubules. NPR-C is located in glomeruli (mainly podocytes), in endothelial cells and PDGFR-β positive cells. To test for a possible regulation of NPRs in kidney diseases, their distribution was studied in adenine nephropathy. Signal intensity of NPR-A and NPR-B mRNA was reduced while their spatial distribution was unaltered compared with healthy kidneys. In contrast, NPR-C mRNA signal was markedly enhanced in cell clusters of myofibroblasts in fibrotic areas of adenine kidneys. In conclusion, the primary renal targets of ANP and BNP are glomerular, vascular, and interstitial cells but not the tubular compartment, while the CNP receptor NPR-B is highly expressed in proximal tubules. Further studies are needed to clarify the function and interplay of this specific receptor expression pattern.
Han X, He Y, Bi GH, Zhang HY, Song R, Liu QR, Egan JM, Gardner EL, Li J, Xi ZX.
PMID: 28951549 | DOI: 10.1038/s41598-017-12399-z
Cannabis can be rewarding or aversive. Cannabis reward is believed to be mediated by activation of cannabinoid CB1 receptors (CB1Rs) on GABAergic neurons that disinhibit dopaminergic neurons in the ventral tegmental area (VTA). However, little is known about the mechanisms underlying cannabis aversion in rodents. In the present study, CB1Rs are found not only on VTA GABAergic neurons, but also on VTA glutamatergic neurons that express vesicular glutamate transporter 2 (VgluT2). We then used Cre-Loxp transgenic technology to selectively delete CB1Rs in VgluT2-expressing glutamatergic neurons (VgluT2-CB1 -/-) and Cre-dependent viral vector to express light-sensitive channelrhodopsin-2 into VTA glutamatergic neurons. We found that photoactivation of VTA glutamatergic neurons produced robust intracranial self-stimulation (ICSS) behavior, which was dose-dependently blocked by DA receptor antagonists, but enhanced by cocaine. In contrast, Δ9-tetrahydrocannabinol (Δ9-THC), the major psychoactive component of cannabis, produced dose-dependent conditioned place aversion and a reduction in the above optical ICSS in VgluT2-cre control mice, but not in VgluT2-CB1 -/- mice. These findings suggest that activation of CB1Rs in VgluT2-expressing glutamate neurons produces aversive effects that might explain why cannabinoid is not rewarding in rodents and might also account for individual differences in the hedonic effects of cannabis in humans.
Cannabinoid CB2 receptors are expressed in glutamate neurons in the red nucleus and functionally modulate motor behavior in mice
Zhang, HY;Shen, H;Gao, M;Ma, Z;Hempel, B;Bi, GH;Gardner, EL;Wu, J;Xi, ZX;
PMID: 33789118 | DOI: 10.1016/j.neuropharm.2021.108538
Cannabinoids produce a number of central nervous system effects via the CB2 receptor (CB2R), including analgesia, antianxiety, anti-reward, hypoactivity and attenuation of opioid-induced respiratory depression. However, the cellular distributions of the CB2Rs in the brain remain unclear. We have reported that CB2Rs are expressed in midbrain dopamine (DA) neurons and functionally regulate DA-mediated behavior(s). Unexpectedly, high densities of CB2-like signaling were also found in a neighboring motor structure - the red nucleus (RN) of the midbrain. In the present study, we systematically explored CB2R expression and function in the RN. Immunohistochemistry and in situ hybridization assays showed high densities of CB2R-immunostaining and mRNA signal in RN magnocellular glutamate neurons in wildtype and CB1-knockout, but not CB2-knockout, mice. Ex vivo electrophysiological recordings in midbrain slices demonstrated that CB2R activation by JWH133 dose-dependently inhibited firing rates of RN magnocellular neurons in wildtype, but not CB2-knockout, mice, while having no effect on RN GABA neurons in transgenic GAD67-GFP reporter mice, suggesting CB2-mediated effects on glutamatergic neurons. In addition, microinjection of JWH133 into the RN produced robust ipsilateral rotations in wildtype, but not CB2-knockout mice, which was blocked by pretreatment with either a CB2 or DA D1 or D2 receptor antagonist, suggesting a DA-dependent effect. Finally, fluorescent tract tracing revealed glutamatergic projections from the RN to multiple brain areas including the ventral tegmental area, nucleus accumbens, thalamus, and cerebellum. These findings suggest that CB2Rs in RN glutamate neurons functionally modulate motor activity, and therefore, constitute a new target in cannabis-based medication development for motor disorders.
Moll S, Yasui Y, Abed A, Murata T, Shimada H, Maeda A, Fukushima N, Kanamori M, Uhles S, Badi L, Cagarelli T, Formentini I, Drawnel F, Georges G, Bergauer T, Gasser R, Bonfil RD, Fridman R, Richter H, Funk J, Moeller MJ, Chatziantoniou C, Prunotto M.
PMID: 29859097 | DOI: 10.1186/s12967-018-1524-5
Abstract
BACKGROUND:
Discoidin domain receptor 1 (DDR1) is a collagen-activated receptor tyrosine kinase extensively implicated in diseases such as cancer, atherosclerosis and fibrosis. Multiple preclinical studies, performed using either a gene deletion or a gene silencing approaches, have shown this receptor being a major driver target of fibrosis and glomerulosclerosis.
METHODS:
The present study investigated the role and relevance of DDR1 in human crescentic glomerulonephritis (GN). Detailed DDR1 expression was first characterized in detail in human GN biopsies using a novel selective anti-DDR1 antibody using immunohistochemistry. Subsequently the protective role of DDR1 was investigated using a highly selective, novel, small molecule inhibitor in a nephrotoxic serum (NTS) GN model in a prophylactic regime and in the NEP25 GN mouse model using a therapeutic intervention regime.
RESULTS:
DDR1 expression was shown to be mainly limited to renal epithelium. In humans, DDR1 is highly induced in injured podocytes, in bridging cells expressing both parietal epithelial cell (PEC) and podocyte markers and in a subset of PECs forming the cellular crescents in human GN. Pharmacological inhibition of DDR1 in NTS improved both renal function and histological parameters. These results, obtained using a prophylactic regime, were confirmed in the NEP25 GN mouse model using a therapeutic intervention regime. Gene expression analysis of NTS showed that pharmacological blockade of DDR1 specifically reverted fibrotic and inflammatory gene networks and modulated expression of the glomerular cell gene signature, further validating DDR1 as a major mediator of cell fate in podocytes and PECs.
CONCLUSIONS:
Together, these results suggest that DDR1 inhibition might be an attractive and promising pharmacological intervention for the treatment of GN, predominantly by targeting the renal epithelium.
Zalachoras, I;Astori, S;Meijer, M;Grosse, J;Zanoletti, O;de Suduiraut, IG;Deussing, JM;Sandi, C;
PMID: 35319997 | DOI: 10.1126/sciadv.abj9019
Individuals frequently differ in their behavioral and cognitive responses to stress. However, whether motivation is differently affected by acute stress in different individuals remains to be established. By exploiting natural variation in trait anxiety in outbred Wistar rats, we show that acute stress facilitates effort-related motivation in low anxious animals, while dampening effort in high anxious ones. This model allowed us to address the mechanisms underlying acute stress-induced differences in motivated behavior. We show that CRHR1 expression levels in dopamine neurons of the ventral tegmental area (VTA)-a neuronal type implicated in the regulation of motivation-depend on animals' anxiety, and these differences in CRHR1 expression levels explain the divergent effects of stress on both effortful behavior and the functioning of mesolimbic DA neurons. These findings highlight CRHR1 in VTA DA neurons-whose levels vary with individuals' anxiety-as a switching mechanism determining whether acute stress facilitates or dampens motivation.
Porcu, A;Nilsson, A;Booreddy, S;Barnes, SA;Welsh, DK;Dulcis, D;
PMID: 36054362 | DOI: 10.1126/sciadv.abn9867
Seasonal changes in day length (photoperiod) affect numerous physiological functions. The suprachiasmatic nucleus (SCN)-paraventricular nucleus (PVN) axis plays a key role in processing photoperiod-related information. Seasonal variations in SCN and PVN neurotransmitter expression have been observed in humans and animal models. However, the molecular mechanisms by which the SCN-PVN network responds to altered photoperiod is unknown. Here, we show in mice that neuromedin S (NMS) and vasoactive intestinal polypeptide (VIP) neurons in the SCN display photoperiod-induced neurotransmitter plasticity. In vivo recording of calcium dynamics revealed that NMS neurons alter PVN network activity in response to winter-like photoperiod. Chronic manipulation of NMS neurons is sufficient to induce neurotransmitter switching in PVN neurons and affects locomotor activity. Our findings reveal previously unidentified molecular adaptations of the SCN-PVN network in response to seasonality and the role for NMS neurons in adjusting hypothalamic function to day length via a coordinated multisynaptic neurotransmitter switching affecting behavior.
