Shi Z, Cassaglia PA, Pelletier NE, Brooks VL.
PMID: PMID: 30628058 | DOI: DOI:10.1113/JP277517
KEY POINTS: ICV insulin increased SNA and baroreflex control of SNA and HR dramatically more in obese male rats; in obese females, the responses were abolished. In obese males, the enhanced LSNA responses were associated with reduced tonic inhibition of LSNA by NPY in the PVN. Yet, PVN NPY injection decreased LSNA similarly in OP/OR/CON rats. Collectively, these results suggest that NPY inputs were decreased. In obese females, NPY inhibition in the PVN was maintained. Moreover, NPY neurons in the ArcN became resistant to the inhibitory effects of insulin. A HFD did not alter arcuate NPY neuronal InsR expression in males or females. Obesity-induced "selective sensitization" of the brain to the sympathoexcitatory effects of insulin and leptin may contribute to elevated basal SNA, and therefore hypertension development, in males with obesity. These data may explain in part why obesity increases SNA less in women compared to men. ABSTRACT: Obesity increases sympathetic nerve activity (SNA) in men, but not women; however, the mechanisms are unknown. We tested if intracerebroventricular insulin infusion increases SNA more in obese male than female rats and if sex differences are mediated by changes in tonic inhibition of SNA by Neuropeptide Y (NPY) in the paraventricular nucleus (PVN). When consuming a high fat diet, obesity prone (OP) rats accrued excess fat, whereas obesity resistant (OR) rats maintained adiposity as in rats eating a control (CON) diet. Insulin increased lumbar SNA (LSNA) similarly in CON/OR males and females under urethane-anesthesia. The LSNA response was magnified in OP males, but abolished in OP females. In males, blockade of PVN NPY Y1 receptors with BIBO3304 increased LSNA in CON/OR rats, but not OP rats. Yet, PVN nanoinjections of NPY decreased LSNA similarly between groups. Thus, tonic PVN NPY inhibition of LSNA may be lost in obese males, due to a decrease in NPY inputs. In contrast, in females, PVN BIBO3304 increased LSNA similarly in OP, OR and CON rats. After insulin, PVN BIBO3304 failed to increase LSNA in CON/OR females, but increased LSNA in OP females, suggesting that with obesity NPY neurons become resistant to the inhibitory effects of insulin. These sex differences were not associated with changes in arcuate NPY neuronal insulin receptor expression. Collectively, these data reveal a marked sex difference in the impact of obesity on insulin's sympathoexcitatory actions and implicate sexually dimorphic changes in NPY inhibition of SNA in the PVN as one mechanism.
Zhang N, Zhang HY, Bi SA, Moran TH and Bi S
PMID: 30902570 | DOI: 10.1016/j.neulet.2019.03.030
Thyrotropin-releasing hormone (TRH) plays an important role in the regulation of energy balance. While the regulation of TRH in the paraventricular nucleus (PVN) in response to changes of energy balance has been well studied, how TRH is regulated in the dorsomedial hypothalamus (DMH) in maintaining energy homeostasis remains unclear. Here, we assessed the effects of food restriction and exercise on hypothalamic Trh expression using Otsuka Long-Evens Tokushima Fatty (OLETF) rats. Sedentary ad lib fed OLETF rats (OLETF-SED) became hyperphagic and obese. These alterations were prevented in OLETF rats with running wheel access (OLETF-RW) or food restriction in which their food was pair-fed (OLETF-PF) to the intake of lean control rats (LETO-SED). Evaluation of hypothalamic gene expression revealed that Trh mRNA expression was increased in the PVN of OLETF-SED rats and normalized in OLETF-RW and OLETF-PF rats compared to LETO-SED rats. In contrast, the expression of Trh in the DMH was decreased in OLETF-SED rats relative to LETO-SED rats. This alteration was reversed in OLETF-RW rats as seen in LETO-SED rats, but food restriction resulted in a significant increase in DMH Trh expression in OLETF-PF rats compared to LETO-SED rats. Strikingly, while Trh mRNA expression was decreased in the PVN of intact rats in response to acute food deprivation, food deprivation resulted in increased expression of Trh in the DMH. Together, these results demonstrate the differential regulation of Trh expression in the PVN and DMH in OLETF rats and suggest that DMH TRH also contributes to hypothalamic regulation of energy balance.
Asian Pac J Cancer Prev. 2015;16(11):4549-53.
Raluca BA, Cimpean AM, Cioca A, Cretu O, Mederle O, Ciolofan A, Gaje P, Raica M.
PMID: 26107202
Abstract BACKGROUND: . Colorectal carcinoma (CRC) is one of the major causes of cancer death worldwide. Data from the literature indicate differences between the proliferation rate of endothelial cells relative to the morphology growth type, possibly due to origin of specimens (autopsy material, surgery fragments) or quantification methods. Vascular endothelial growth factor (VEGF) is a factor that stimulates the proliferation of endothelial cells. It is expressed in more than 90% of cases of metastatic CRC. AIM: The aim of this study was to evaluate the endothelial cell proliferation and VEGF expression in primary tumors and corresponding liver metastases. MATERIALS AND METHODS: Our study included 24 recent biopsies of primary tumors and corresponding liver metastases of CRC cases. CD34/ Ki67 double immunostaining and RNA scope assay for VEGF were performed. RESULTS: In the primary tumors analysis of VEGFmRNA expression indicated no significant correlation with differentiation grade, proliferative and non-proliferative vessels in the intratumoral and peritumoral areas. In contrast, in the corresponding liver metastases, VEGFmRNA expression significantly correlated with the total number of non- proliferative vessels and total number of vessels. CD34/ Ki67 double immunostaining in the cases with poorly differentiated carcinoma indicated a high number of proliferating endothelial cells in the peritumoral area and a low number in the intratumoral area for the primary tumor. Moderately differentiated carcinomas of colon showed no proliferating endothelial cells in the intratumoral area in half of the cases included in the study, for both, primary tumor and liver metastasis. In well differentiated CRCs, in primary tumors, a high proliferation rate of endothelial cells in the intratumoral area and a lower proliferation rate in the peritumoral area were found. A low value was found in corresponding liver metastasis. CONCLUSIONS: The absence of proliferative endothelial cells in half of the cases for the primary tumors and liver metastases in moderately differentiated carcinoma suggest a vascular mimicry phenomenon. The mismatch between the total number of vessels and endothelial proliferation in primary tumors indicate that a functional vascular network is already formed or the existence of some mechanisms influenced by other angiogenic factors.
Frontiers in molecular biosciences
Nabi, R;Musarrat, F;Menk P Lima, JC;Langohr, IM;Chouljenko, VN;Kousoulas, KG;
PMID: 37388243 | DOI: 10.3389/fmolb.2023.1199068
Introduction: Oncolytic viruses (OVs) provide new modalities for cancer therapy either alone or in combination with synergistic immunotherapies and/or chemotherapeutics. Engineered Herpes Simplex Virus Type-1 (HSV-1) has shown strong promise for the treatment of various cancers in experimental animal models as well as in human patients, with some virus strains licensed to treat human melanoma and gliomas. In the present study we evaluated the efficacy of mutant HSV-1 (VC2) in a late stage, highly metastatic 4T1 murine syngeneic. Method: VC2 was constructed VC2 using double red recombination technology. For in-vivo efficacy we utilized a late stage 4T1 syngeneic and immunocompetent BALB/cJ mouse model breast cancer model which exhibits efficient metastasis to the lung and other organs. Results: VC2 replicated efficiently in 4T1 cells and in cell culture, achieving titers similar to those in African monkey kidney (Vero) cells. Intra-tumor treatment with VC2 did not appreciably reduce average primary tumor sizes but a significant reduction of lung metastasis was noted in mice treated intratumorally with VC2, but not with ultraviolet-inactivated VC2. This reduction of metastasis was associated with increased T cell infiltration comprised of CD4+ and CD4+CD8+ double-positive T cells. Characterization of purified tumor infiltrating T cells revealed a significant improvement in their proliferation ability compared to controls. In addition, significant T cell infiltration was observed in the metastatic nodules associated with reduction of pro-tumor PD-L1 and VEGF gene transcription. Conclusion: These results show that VC2 therapy can improve anti-tumor response associated with a better control of tumor metastasis. improve T cell responses and reduce pro-tumor biomarker gene transcription. VC2 holds promise for further development as an oncolytic and immunotherapeutic approach to treat breast and other cancers.
The Journal of neuroscience : the official journal of the Society for Neuroscience
Ambler, M;Hitrec, T;Wilson, A;Cerri, M;Pickering, A;
PMID: 35440490 | DOI: 10.1523/JNEUROSCI.2102-21.2022
Torpor is a naturally occurring, hypometabolic, hypothermic state engaged by a wide range of animals in response to imbalance between the supply and demand for nutrients. Recent work has identified some of the key neuronal populations involved in daily torpor induction in mice, in particular projections from the preoptic area of the hypothalamus (POA) to the dorsomedial hypothalamus (DMH). The DMH plays a role in thermoregulation, control of energy expenditure, and circadian rhythms, making it well positioned to contribute to the expression of torpor. We used activity dependent genetic TRAPing techniques to target DMH neurons that were active during natural torpor bouts in female mice. Chemogenetic reactivation of torpor-TRAPed DMH neurons in calorie-restricted mice promoted torpor, resulting in longer and deeper torpor bouts. Chemogenetic inhibition of torpor-TRAPed DMH neurons did not block torpor entry, suggesting a modulatory role for the DMH in the control of torpor. This work adds to the evidence that the POA and the DMH form part of a circuit within the mouse hypothalamus that controls entry into daily torpor.SIGNIFICANCEDaily heterotherms such as mice employ torpor to cope with environments in which the supply of metabolic fuel is not sufficient for the maintenance of normothermia. Daily torpor involves reductions in body temperature, as well as active suppression of heart rate and metabolism. How the central nervous system controls this profound deviation from normal homeostasis is not known, but a projection from the preoptic area to the dorsomedial hypothalamus has recently been implicated. We demonstrate that the dorsomedial hypothalamus contains neurons that are active during torpor. Activity in these neurons promotes torpor entry and maintenance, but their activation alone does not appear to be sufficient for torpor entry.
Journal of cellular physiology
Zhang, CL;Lin, YZ;Wu, Q;Yan, C;Wong, MW;Zeng, F;Zhu, P;Bowes, K;Lee, K;Zhang, X;Song, ZY;Lin, S;Shi, YC;
PMID: 35312067 | DOI: 10.1002/jcp.30719
Chronic high salt intake is one of the leading causes of hypertension. Salt activates the release of the key neurotransmitters in the hypothalamus such as vasopressin to increase blood pressure, and neuropepetide Y (NPY) has been implicated in the modulation of vasopressin levels. NPY in the hypothalamic arcuate nucleus (Arc) is best known for its control in appetite and energy homeostasis, but it is unclear whether it is also involved in the development of salt-induced hypertension. Here, we demonstrate that wild-type mice given 2% NaCl salt water for 8 weeks developed hypertension which was associated with marked downregulation of NPY expression in the hypothalamic Arc as demonstrated in NPY-GFP reporter mice as well as by in situ hybridization analysis. Furthermore, salt intake activates neurons in the hypothalamic paraventricular nucleus (PVN) where mRNA expression of brain-derived neurotrophic factor (BDNF) and vasopressin was found to be upregulated, leading to elevated serum vasopressin levels. This finding suggests an inverse correlation between the Arc NPY level and expression of vasopressin and BDNF in the PVN. Specific restoration of NPY by injecting AAV-Cre recombinase into the Arc only of the NPY-targeted mutant mice carrying a loxP-flanked STOP cassette reversed effects of salt intake on vasopressin and BDNF expression, leading to a normalization of salt-dependent blood pressure. In summary, our study uncovers an important Arc NPY-originated neuronal circuitry that could sense and respond to peripheral electrolyte signals and thereby regulate hypertension via vasopressin and BDNF in the PVN.
Harris NA, Isaac AT, Günther A, Merkel K, Melchior J, Xu M, Eguakun E, Perez R, Nabit BP, Flavin S, Gilsbach R, Shonesy B, Hein L, Abel T, Baumann A, Matthews R, Centanni SW, Winder DG.
PMID: 30150361 | DOI: 10.1523/JNEUROSCI.0963-18.2018
Stress is a precipitating agent in neuropsychiatric disease and initiates relapse to drug-seeking behavior in addicted patients. Targeting the stress system in protracted abstinence from drugs of abuse with anxiolytics may be an effective treatment modality for substance use disorders. α2A-adrenergic receptors (α2A-ARs) in extended amygdala structures play key roles in dampening stress responses. Contrary to early thinking, α2A-ARs are expressed at non-noradrenergic sites in the brain. These non-noradrenergic α2A-ARs play important roles in stress-responses, but their cellular mechanisms of action are unclear. In humans, the α2A-AR agonist guanfacine reduces overall craving and uncouples craving from stress yet minimally affects relapse, potentially due to competing actions in the brain. Here we show that heteroceptor α2A-ARs postsynaptically enhance dorsal BNST (dBNST) neuronal activity in mice of both sexes. This effect is mediated by hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels, as inhibition of these channels is necessary and sufficient for excitatory actions. Finally, this excitatory action is mimicked by clozapine-N-oxide activation of the Gi-coupled DREADD hM4Di in dBNST neurons, and its activation elicits anxiety-like behavior in the elevated plus maze. Together, this data provides a framework for elucidating cell-specific actions of GPCR signaling and provides a potential mechanism whereby competing anxiogenic and anxiolytic actions of guanfacine may affect its clinical utility in the treatment of addiction.SIGNIFICANCE STATEMENTStress impacts the development of neuropsychiatric disorders including anxiety and addiction. Guanfacine is an α2A-adrenergic receptor (α2A-AR) agonist with actions in the bed nucleus of the stria terminalis (BNST) that produces antidepressant actions and uncouples stress from reward-related behaviors. Here we show that guanfacine increases dBNST neuronal activity through actions at postsynaptic α2A-ARs via a mechanism that involves hyperpolarization-activated cyclic nucleotide gated cation (HCN) channels. This action is mimicked by activation of the designer receptor hM4Di expressed in the BNST, which also induces anxiety-like behaviors. Together, these data suggest 1) that postsynaptic α2A-ARs in BNST have excitatory actions on BNST neurons, and 2) these actions can be phenocopied by the so-called "inhibitory" DREADDs, suggesting care must be taken regarding interpretation of data obtained with these tools.
Distinct Cellular Profiles of Hif1a and Vegf mRNA Localization in Microglia, Astrocytes and Neurons during a Period of Vascular Maturation in the Auditory Brainstem of Neonate Rats
Chang, D;Brown, Q;Tsui, G;He, Y;Liu, J;Shi, L;Rodríguez-Contreras, A;
| DOI: 10.3390/brainsci11070944
Defining the relationship between vascular development and the expression of hypoxia-inducible factors (Hifs) and vascular endothelial growth factor (Vegf) in the auditory brainstem is important to understand how tissue hypoxia caused by oxygen shortage contributes to sensory deficits in neonates. In this study, we used histology, molecular labeling, confocal microscopy and 3D image processing methods to test the hypothesis that significant maturation of the vascular bed in the medial nucleus of the trapezoid body (MNTB) occurs during the postnatal period that precedes hearing onset. Isolectin-B4 histochemistry experiments suggested that the MNTB vasculature becomes more elaborate between P5 and P10. When combined with a cell proliferation marker and immunohistochemistry, we found that vascular growth coincides with a switch in the localization of proliferating cells to perivascular locations, and an increase in the density of microglia within the MNTB. Furthermore, microglia were identified as perivascular cells with proliferative activity during the period of vascular maturation. Lastly, combined in situ hybridization and immunohistochemistry experiments showed distinct profiles of Hif1a and Vegf mRNA localization in microglia, astrocytes and MNTB principal neurons. These results suggest that different cells of the neuro-glio-vascular unit are likely targets of hypoxic insult in the auditory brainstem of neonate rats.
Journal of molecular medicine (Berlin, Germany)
Sych, K;Nold, SP;Pfeilschifter, J;Vutukuri, R;Meisterknecht, J;Wittig, I;Frank, S;Goren, I;
PMID: 36633604 | DOI: 10.1007/s00109-022-02280-6
An injured skin is rapidly restored in a manner of wound healing. We have previously shown that intact insulin signaling and glucose uptake are fundamental to proper wound closure. Consequently, under exacerbated inflammation, compromised insulin action and glucose uptake lead to impaired healing. However, in spite of the increased attention to cell metabolism during tissue regeneration, metabolic mediators that govern cellular and physiological processes throughout skin repair remained largely elusive. Through assessment of mRNA using real-time PCR and protein blot analysis, we report that healing of cutaneous wounds comprise a boosted expression of genes involved in glycolysis, oxidative phosphorylation, pentose phosphate shunt, and glutamine anaplerosis. We further focused on the functional role of pyruvate kinase M (PKM) isoenzymes that catalyze the final and rate-limiting step of glycolysis. Whereas the expression of the metabolic constitutively active Pkm1 isozyme remained almost unchanged, Pkm2 is augmented during the inflammatory phase of healing. The immunohistochemistry and RNA in situ hybridization analysis showed a confined Pkm2 expression to keratinocytes of the hyperproliferative epithelium and, to a lesser extent, infiltrating neutrophils and monocytes as well as later on in macrophages. Notably, the expression of Pkm2 in keratinocytes facing the wound bed side colocalized with VEGF expression. The in vitro knockdown of PKM2 in HaCaT keratinocytes using small interfering (si) RNA confirmed an acute role for PKM2 in facilitating the complete induction of VEGF mRNA and protein expression in keratinocytes; this function is mainly HIF-1α independent. KEY MESSAGES: • Wound healing involves activation of glycolysis, oxidative phosphorylation, pentos-phosphate shunt, and replenishment of tri-carboxylic acid (TCA) cycle through glutamine anaplerosis. • The pyruvate kinase M2 (PKM2) isoform is upregulated during the inflammatory phase of cutaneous healing, mainly in keratinocytes of hyperproliferative epithelia. • In vivo, the expression of VEGF in wound keratinocytes is colocalized with PKM2. • PKM2 is required for full induction of VEGF in HaCaT keratinocytes in vitro.
Shi, Z;Stornetta, DS;Stornetta, RL;Brooks, VL;
PMID: 34937769 | DOI: 10.1523/ENEURO.0404-21.2021
The arcuate nucleus (ArcN) is an integrative hub for the regulation of energy balance, reproduction, and arterial pressure (AP), all of which are influenced by Angiotensin II (AngII); however, the cellular mechanisms and downstream neurocircuitry are unclear. Here we show that ArcN AngII increases AP in female rats via two phases, both of which are mediated via activation of AngII type 1 receptors (AT1aR): initial vasopressin-induced vasoconstriction, followed by slowly developing increases in sympathetic nerve activity (SNA) and heart rate (HR). In male rats, ArcN AngII evoked a similarly slow increase in SNA, but the initial pressor response was variable. In females, the effects of ArcN AngII varied during the estrus cycle, with significant increases in SNA, HR, and AP occurring during diestrus and estrus, but only increased AP during proestrus. Pregnancy markedly increased the expression of AT1aR in the ArcN with parallel substantial AngII-induced increases in SNA and MAP. In both sexes, the sympathoexcitation relied on suppression of tonic ArcN sympathoinhibitory Neuropeptide Y inputs, and activation of pro-opiomelanocortin (POMC) projections, to the paraventricular nucleus (PVN). Few or no NPY or POMC neurons expressed the AT1aR, suggesting that AngII increases AP and SNA at least in part indirectly via local interneurons, which express tyrosine hydroxylase (TH) and VGat (i.e. GABAergic). ArcN TH neurons release GABA locally, and central AT1aR and TH neurons mediate stress responses; therefore, we propose that TH AT1aR neurons are well situated to locally coordinate the regulation of multiple modalities within the ArcN in response to stress.SIGNIFICANCEThe arcuate nucleus (ArcN) is an integrative hub for the regulation of energy balance, reproduction, and arterial pressure (AP), all of which are influenced by Angiotensin II (AngII). Here we show that ArcN AngII activates AT1aR to increase AP in male and female rats by slowly increasing sympathetic nerve activity. In females, ArcN AngII also evoked an initial pressor response mediated by vasopressin-induced vasoconstriction. Pregnant and estrus females responded more than males, in association with higher ArcN AT1aR expression. AT1aR were identified in ArcN interneurons that express tyrosine hydroxylase (TH) and GABA. Since brain AT1aR and TH mediate stress responses, ArcN AT1aR TH neurons are well situated to locally coordinate autonomic, hormonal, and behavioral responses to stress.
Lee SJ, Sanchez-Watts G, Krieger JP, Pignalosa A, Norell PN, Cortella A, Pettersen KG, Vrdoljak D, Hayes MR, Kanoski S, Langhans W, Watts AG.
PMID: - | DOI: 10.1016/j.molmet.2018.03.008
Abstract
Objective
Glucagon-like peptide-1 (GLP-1) neurons in the hindbrain densely innervate the dorsomedial hypothalamus (DMH), a nucleus strongly implicated in body weight regulation and the sympathetic control of brown adipose tissue (BAT) thermogenesis. Therefore, DMH GLP-1 receptors (GLP-1R) are well placed to regulate energy balance by controlling sympathetic outflow and BAT function.
Methods
We investigate this possibility in adult male rats by using direct administration of GLP-1 (0.5 ug) into the DMH, knocking down DMH GLP-1R mRNA with viral-mediated RNA interference, and by examining the neurochemical phenotype of GLP-1R expressing cells in the DMH using in situ hybridization.
Results
GLP-1 administered into the DMH increased BAT thermogenesis and hepatic triglyceride (TG) mobilization. On the other hand, Glp1r knockdown (KD) in the DMH increased body weight gain and adiposity, with a concomitant reduction in energy expenditure (EE), BAT temperature, and uncoupling protein 1 (UCP1) expression. Moreover, DMH Glp1r KD induced hepatic steatosis, increased plasma TG, and elevated liver specific de-novo lipogenesis, effects that collectively contributed to insulin resistance. Interestingly, DMH Glp1r KD increased neuropeptide Y (NPY) mRNA expression in the DMH. GLP-1R mRNA in the DMH, however, was found in GABAergic not NPY neurons, consistent with a GLP-1R-dependent inhibition of NPY neurons that is mediated by local GABAergic neurons. Finally, DMH Glp1r KD attenuated the anorexigenic effects of the GLP-1R agonist exendin-4, highlighting an important role of DMH GLP-1R signaling in GLP-1-based therapies.
Conclusions
Collectively, our data show that DMH GLP-1R signaling plays a key role for BAT thermogenesis and adiposity.
Chen YW, Das M, Oyarzabal EA, Cheng Q, Plummer NW, Smith KG, Jones GK, Malawsky D, Yakel JL, Shih YI, Jensen P.
PMID: 30214043 | DOI: 10.1038/s41380-018-0245-8
Noradrenergic signaling plays a well-established role in promoting the stress response. Here we identify a subpopulation of noradrenergic neurons, defined by developmental expression of Hoxb1, that has a unique role in modulating stress-related behavior. Using an intersectional chemogenetic strategy, in combination with behavioral and physiological analyses, we show that activation of Hoxb1-noradrenergic (Hoxb1-NE) neurons decreases anxiety-like behavior and promotes an active coping strategy in response to acute stressors. In addition, we use cerebral blood volume-weighted functional magnetic resonance imaging to show that chemoactivation of Hoxb1-NE neurons results in reduced activity in stress-related brain regions, including the bed nucleus of the stria terminalis, amygdala, and locus coeruleus. Thus, the actions of Hoxb1-NE neurons are distinct from the well-documented functions of the locus coeruleus in promoting the stress response, demonstrating that the noradrenergic system contains multiple functionally distinct subpopulations.
