RESUMO
One feature of high-fat diet-induced neurodegeneration in the hypothalamus is an increased level of palmitate, which is associated with endoplasmic reticulum (ER) stress, loss of CoxIV, mitochondrial fragmentation, and decreased abundance of MC4R. To determine whether antidiabetic drugs protect against ER and/or mitochondrial dysfunction by lipid stress, hypothalamic neurons derived from pre-adult mice and neuronal Neuro2A cells were exposed to elevated palmitate. In the hypothalamic neurons, palmitate exposure increased expression of ER resident proteins, including that of SERCA2, indicating ER stress. Liraglutide reverted such altered ER proteostasis, while metformin only normalized SERCA2 expression. In Neuro2A cells liraglutide, but not metformin, also blunted dilation of the ER induced by palmitate treatment, and enhanced abundance and expression of MC4R at the cell surface. Thus, liraglutide counteracts, more effectively than metformin, altered ER proteostasis, morphology, and folding capacity in neurons exposed to fat. In palmitate-treated hypothalamic neurons, mitochondrial fragmentation took place together with loss of CoxIV and decreased mitochondrial membrane potential (MMP). Metformin, but not liraglutide, reverted mitochondrial fragmentation, and both liraglutide and metformin did not protect against either loss of CoxIV abundance or MMP. Thus, ER recovery from lipid stress can take place in hypothalamic neurons in the absence of recovered mitochondrial homeostasis.
Assuntos
Liraglutida , Metformina , Animais , Camundongos , Liraglutida/farmacologia , Palmitatos/farmacologia , Palmitatos/metabolismo , Estresse do Retículo Endoplasmático , Hipotálamo/metabolismo , Neurônios/metabolismo , Metformina/farmacologia , Metformina/metabolismo , Mitocôndrias/metabolismoRESUMO
The melanocortin-4 receptor (MC4R) is a G protein-coupled receptor (GPCR) that is expressed in several brain locations encompassing the hypothalamus and the brainstem, where the receptor controls several body functions, including metabolism. In a well-defined pathway to decrease appetite, hypothalamic proopiomelanocortin (POMC) neurons localized in the arcuate nucleus (Arc) project to MC4R neurons in the paraventricular nuclei (PVN) to release the natural MC4R agonist α-melanocyte-stimulating hormone (α-MSH). Arc neurons also project excitatory glutamatergic fibers to the MC4R neurons in the PVN for a fast synaptic transmission to regulate a satiety pathway potentiated by α-MSH. By using super-resolution microscopy, we found that in hypothalamic neurons in a primary culture, postsynaptic density protein 95 (PSD95) colocalizes with GluN1, a subunit of the ionotropic N-methyl-D-aspartate receptor (NMDAR). Thus, hypothalamic neurons form excitatory postsynaptic specializations. To study the MC4R distribution at these sites, tagged HA-MC4R under the synapsin promoter was expressed in neurons by adeno-associated virus (AAV) gene transduction. HA-MC4R immunofluorescence peaked at the center and in proximity to the PSD95- and NMDAR-expressing sites. These data provide morphological evidence that MC4R localizes together with glutamate receptors at postsynaptic and peri-postsynaptic sites.
Assuntos
Hipotálamo , Neurônios , Receptor Tipo 4 de Melanocortina , Animais , Receptor Tipo 4 de Melanocortina/metabolismo , Receptor Tipo 4 de Melanocortina/genética , Neurônios/metabolismo , Hipotálamo/metabolismo , Hipotálamo/citologia , Camundongos , Sinapses/metabolismo , Proteína 4 Homóloga a Disks-Large/metabolismo , Células Cultivadas , Receptores de N-Metil-D-Aspartato/metabolismoRESUMO
In the melanocortin pathway, melanocortin-4 receptor (MC4R) functions to control energy homeostasis. MC4R is expressed in a sub-population of Sim1 neurons (Sim1/MC4R neurons) and functions in hypothalamic paraventricular nuclei (PVN) to control food intake. Mapping sites of hypothalamic injury in obesity is essential to counteract the disease. In the PVN of male and female mice with diet-induced obesity (DIO) there is neuronal loss. However, the existing subpopulation of PVN Sim1/MC4R neurons is unchanged, but has a loss of mitochondria and MC4R protein. In mice of both sexes with DIO, dietary intervention to re-establish normal weight restores abundance of MC4R protein in Sim1/MC4R neurons and neurogenesis in the PVN. However, the number of non-Sim1/MC4R neurons in the PVN continues to remain decreased. Selective survival and recovery of Sim1/MC4R neurons after DIO suggests these neurons as preferential target to restore energy homeostasis and of therapy against obesity.
RESUMO
Genetic obesity increases in liver phosphatidylcholine (PC)/phosphatidylethanolamine (PE) ratio, inducing endoplasmic reticulum (ER) stress without concomitant increase of ER chaperones. Here, it is found that exposing mice to a palm oil-based high fat (HF) diet induced obesity, loss of liver PE, and loss of the ER chaperone Grp78/BiP in pericentral hepatocytes. In Hepa1-6 cells treated with elevated concentration of palmitate to model lipid stress, Grp78/BiP mRNA was increased, indicating onset of stress-induced Unfolded Protein Response (UPR), but Grp78/BiP protein abundance was nevertheless decreased. Exposure to elevated palmitate also induced in hepatoma cells decreased membrane glycosylation, nuclear translocation of pro-apoptotic C/EBP-homologous-protein-10 (CHOP), expansion of ER-derived quality control compartment (ERQC), loss of mitochondrial membrane potential (MMP), and decreased oxidative phosphorylation. When PE was delivered to Hepa1-6 cells exposed to elevated palmitate, effects by elevated palmitate to decrease Grp78/BiP protein abundance and suppress membrane glycosylation were blunted. Delivery of PE to Hepa1-6 cells treated with elevated palmitate also blunted expansion of ERQC, decreased nuclear translocation of CHOP and lowered abundance of reactive oxygen species (ROS). Instead, delivery of the chemical chaperone 4-phenyl-butyrate (PBA) to Hepa1-6 cells treated with elevated palmitate, while increasing abundance of Grp78/BiP protein and restoring membrane glycosylation, also increased ERQC, expression and nuclear translocation of CHOP, non-mitochondrial oxygen consumption, and generation of ROS. Data indicate that delivery of PE to hepatoma cells under lipid stress recovers cell function by targeting the secretory pathway and by blunting pro-apoptotic branches of the UPR.