Neuroanatomical characterization of the G protein-coupled receptor activity evoked by galanin-related ligands

Galanin neuropeptide is distributed throughout the mammalian nervous system modulating a plethora of diverse physiological functions, including nociception, cognition and neuroendocrine regulation. The regulation of the galaninergic system is an interesting approach for the treatment of different diseases associated to those systems. Nevertheless, the pharmacological selectivity and activities of some galanin receptor (GalR) ligands are still in discussion and seem to depend on the dose, the receptor subtype and the second messengers to which they are coupled at different brain areas. The activity of different GalR ligands on Gi/o proteins, was evaluated by the guanosine 5′-(γ-[35S]thio)triphosphate ([35S]GTPγS) autoradiography in vitro assay applied to rat brain tissue slices in the presence of galanin, M15, M35, M40, gal(2−11) or galnon. The enhancement of the [35S]GTPγS binding induced by the chimerical peptides M15, M35 and M40 was similar to that produced by Gal in those brain areas showing the highest stimulations, such as dorsal part of the olfactory nucleus and ventral subiculum. In contrast to these peptides, using gal(2−11) no effect was measured on Gi/o protein coupling in areas of the rat brain with high GalR1 density such as posterior hypothalamic nucleus and amygdala, indicating low selectivity for GalR1 receptors. The effects evoked by the non-peptide ligand, galnon, were different from those induced by galanin, behaving as agonist or antagonist depending on the brain area, but the stimulations were always blocked by M35. Thus, the activity of most used GalR ligands on Gi/o protein mediated signalling is complex and depends on the brain area. More selective and potent GalR ligands are necessary to develop new treatments aimed to modulate the galaninergic system.Supported by grants from the regional Basque Government IT1454–22 awarded to the "Neurochemistry and Neurodegeneration" consolidated research group and ISCIII Spanish Ministry for Health PI20/00153 and co-funded by the European Union (ERDF "A way to make Europe"). Technical and human support provided by General Research Services SGIker [University of the Basque Country (UPV/EHU)]

Type-1 cannabinoid receptor activity during Alzheimer's disease progression

The activity of CB1 cannabinoid receptors was studied in postmortem brain samples of Alzheimer's disease (AD) patients during clinical deterioration. CB1 activity was higher at earlier AD stages in limited hippocampal areas and internal layers of frontal cortex, but a decrease was observed at the advanced stages. The pattern of modification appears to indicate initial hyperactivity of the endocannabinoid system in brain areas that lack classical histopathological markers at earlier stages of AD, indicating an attempt to compensate for the initial synaptic impairment, which is then surpassed by disease progression. These results suggest that initial CB1 stimulation might have therapeutic relevance

Microarrays, Enzymatic Assays, and MALDI-MS for Determining Specific Alterations to Mitochondrial Electron Transport Chain Activity, ROS Formation, and Lipid Composition in a Monkey Model of Parkinson’s Disease

Multiple evidences suggest that mitochondrial dysfunction is implicated in the pathogenesis of Parkinson’s disease via the selective cell death of dopaminergic neurons, such as that which occurs after prolonged exposure to the mitochondrial electron transport chain (ETC) complex I inhibitor, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyrine (MPTP). However, the effects of chronic MPTP on the ETC complexes and on enzymes of lipid metabolism have not yet been thoroughly determined. To face these questions, the enzymatic activities of ETC complexes and the lipidomic profile of MPTP-treated non-human primate samples were determined using cell membrane microarrays from different brain areas and tissues. MPTP treatment induced an increase in complex II activity in the olfactory bulb, putamen, caudate, and substantia nigra, where a decrease in complex IV activity was observed. The lipidomic profile was also altered in these areas, with a reduction in the phosphatidylserine (38:1) content being especially relevant. Thus, MPTP treatment not only modulates ETC enzymes, but also seems to alter other mitochondrial enzymes that regulate the lipid metabolism. Moreover, these results show that a combination of cell membrane microarrays, enzymatic assays, and MALDI-MS provides a powerful tool for identifying and validating new therapeutic targets that might accelerate the drug discovery process.This research has been supported by grants from the regional Basque Government ITI1454-22 awarded to the “Neurochemistry and Neurodegeneration” consolidated research group and ISCIII Spanish Ministry for Health PI20/00153 and co-funded by the European Union (ERDF “A way to make Europe”), a grant from the Ministry of Economy and Competitiveness (IPT-2011-1205) and Scholarship Program for the Transition from Educational to Occupational Word (UPV-Basque Government)

Handling and novel object recognition modulate fear response and endocannabinoid signaling in nucleus basalis magnocellularis

[EN] Storage of aversive memories is of utmost importance for survival, allowing animals to avoid upcoming similar stimuli. However, without reinforcement, the learned avoidance response gradually decreases over time. Although the molecular mechanisms controlling this extinction process are not well known, there is evidence that the endocannabinoid system plays a key role through CB1 receptor-mediated modulation of cholinergic signaling. In this study, we measured fear extinction throughout 7 months using naive rats, assessed in passive avoidance (PA) test in a non-reinforced manner. Then, we evaluated the effect of gentle handling and non-aversive novel object recognition test (NORT) on the extinction and expression of fear memories by measuring passive avoidance responses. Neurochemical correlates were analyzed by functional autoradiography for cannabinoid, cholinergic, and dopaminergic receptors. Despite results showing a gradual decrease of passive avoidance response, it did not fully disappear even after 7 months, indicating the robustness of this process. Meanwhile, in rats that received gentle handling or performed NORT after receiving the PA aversive stimulus, extinction occurred within a week. In contrast, gentle handling performed before receiving the aversive stimulus exacerbated fear expression and triggered escape response in PA. The neurochemical analysis showed increased cannabinoid and cholinergic activity in the nucleus basalis magnocellularis (NBM) in rats that had performed only PA, as opposed to rats that received gentle handling before PA. Additionally, a correlation between CB1 mediated-signaling in the NBM and freezing in PA was found, suggesting that the endocannabinoid system might be responsible for modulating fear response induced by aversive memories.Basque Government IT975-16 to the "Neurochemistry and Neurodegeneration" consolidated research group; Instituto de Salud Carlos III, Grant/Award Number: PI20/0015

Spinal Inhibition of GABAB Receptors by the Extracellular Matrix Protein Fibulin-2 in Neuropathic Rats

In the central nervous system, the inhibitory GABAB receptor is the archetype of heterodimeric G protein-coupled receptors (GPCRs). Receptor interaction with partner proteins has emerged as a novel mechanism to alter GPCR signaling in pathophysiological conditions. We propose here that GABAB activity is inhibited through the specific binding of fibulin-2, an extracellular matrix protein, to the B1a subunit in a rat model of neuropathic pain. We demonstrate that fibulin-2 hampers GABAB activation, presumably through decreasing agonist-induced conformational changes. Fibulin-2 regulates the GABAB-mediated presynaptic inhibition of neurotransmitter release and weakens the GABAB-mediated inhibitory effect in neuronal cell culture. In the dorsal spinal cord of neuropathic rats, fibulin-2 is overexpressed and colocalized with B1a. Fibulin-2 may thus interact with presynaptic GABAB receptors, including those on nociceptive afferents. By applying anti-fibulin-2 siRNAin vivo, we enhanced the antinociceptive effect of intrathecal baclofen in neuropathic rats, thus demonstrating that fibulin-2 limits the action of GABAB agonistsin vivo. Taken together, our data provide an example of an endogenous regulation of GABAB receptor by extracellular matrix proteins and demonstrate its functional impact on pathophysiological processes of pain sensitization.This work was funded by the ANR ImNet (ANR-07-NEURO015-01). Imaging was performed on the Bordeaux Imaging Center, member of the FranceBioImaging national infrastructure (ANR-10-INBS-04)

Modulation of Neurolipid Signaling and Specific Lipid Species in the Triple Transgenic Mouse Model of Alzheimer’s Disease

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder and the most common cause of dementia in aging populations. Recently, the regulation of neurolipid-mediated signaling and cerebral lipid species was shown in AD patients. The triple transgenic mouse model (3xTg-AD), harboring βAPPSwe, PS1M146V, and tauP301L transgenes, mimics many critical aspects of AD neuropathology and progressively develops neuropathological markers. Thus, in the present study, 3xTg-AD mice have been used to test the involvement of the neurolipid-based signaling by endocannabinoids (eCB), lysophosphatidic acid (LPA), and sphingosine 1-phosphate (S1P) in relation to the lipid deregulation. [35S]GTPγS autoradiography was used in the presence of specific agonists WIN55,212-2, LPA and CYM5442, to measure the activity mediated by CB1, LPA1, and S1P1 Gi/0 coupled receptors, respectively. Consecutive slides were used to analyze the relative intensities of multiple lipid species by MALDI Mass spectrometry imaging (MSI) with microscopic anatomical resolution. The quantitative analysis of the astrocyte population was performed by immunohistochemistry. CB1 receptor activity was decreased in the amygdala and motor cortex of 3xTg-AD mice, but LPA1 activity was increased in the corpus callosum, motor cortex, hippocampal CA1 area, and striatum. Conversely, S1P1 activity was reduced in hippocampal areas. Moreover, the observed modifications on PC, PA, SM, and PI intensities in different brain areas depend on their fatty acid composition, including decrease of polyunsaturated fatty acid (PUFA) phospholipids and increase of species containing saturated fatty acids (SFA). The regulation of some lipid species in specific brain regions together with the modulation of the eCB, LPA, and S1P signaling in 3xTg-AD mice indicate a neuroprotective adaptation to improve neurotransmission, relieve the myelination dysfunction, and to attenuate astrocyte-mediated neuroinflammation. These results could contribute to identify new therapeutic strategies based on the regulation of the lipid signaling in familial AD patients.This research was funded by the regional Basque Government IT975-16 to the “Neurochemistry and Neurodegeneration” consolidated research group and ISCIII Spanish Ministry for Health PI20/00153

Imaging mass spectrometry (IMS) of cortical lipids from preclinical to severe stages of Alzheimer's disease

Alzheimer's disease (AD) is a progressive neurodegenerative disease affecting millions of patients worldwide. Previous studies have demonstrated alterations in the lipid composition of lipid extracts from plasma and brain samples of AD patients. However, there is no consensus regarding the qualitative and quantitative changes of lipids in brains from AD patients. In addition, the recent developments in imaging mass spectrometry methods are leading to a new stage in the in situ analysis of lipid species in brain tissue slices from human postmortem samples. The present study uses the matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS), permitting the direct anatomical analysis of lipids in postmortem brain sections from AD patients, which are compared with the intensity of the lipid signal in samples from matched subjects with no neurological diseases. The frontal cortex samples from AD patients were classified in three groups based on Braak's histochemical criteria, ranging from non-cognitively impaired patients to those severely affected. The main results indicate a depletion of different sulfatide lipid species from the earliest stages of the disease in both white and gray matter areas of the frontal cortex. Therefore, the decrease in sulfatides in cortical areas could be considered as a marker of the disease, but may also indicate neurochemical modifications related to the pathogenesis of the disease

Lisofosfolipidoen eta Alzheimer gaixotasunaren arteko erlazioa: etorkizuneko itu farmakologikoaren bila

Lipidoak funtzio energetikoaz eta egitura-funtzioez gain deskribatu diren beste funtzioei esker garrantzitsuak bilakatzen ari dira. Funtzio neurotransmisorea edota neuromoduladorea aurkeztu duten lipidoen artean, lisofosfolipidoak aurkitu ditzakegu. Lisofosfolipidoak lipido molekula txiki bioaktiboak, karbonodun kate bakarra eta buru polar bat edukitzeagatik bereizten direnak dira. Lisofosfolipidoen artean, azido lisofosfatidikoa eta esfingosina 1-fosfatoaren egitura eta sistemen funtzioak izan dira hobeto deskribatu direnak. Lisofosfolipidoak zelulaz kanpoko bitartekari aritzen dira berentzat espezifikoak diren G proteinei loturiko hartzaileak aktibatuz. Molekula horien seinaleztapenaren bidez zenbait prozesu neurokimiko modulatzen dira, adibidez, neuromodulazioa eta neuroinflamazioa. Gainera, ikasketarekin eta oroimenarekin erlazioa erakutsi dute. Horren haritik, orain arte ondoen deskribatutako lisofosfolipidoen sistemak, azido lisofosfatidikoa eta esfingosina l-fosfatoa, hain zuzen ere, asaldatuta daude Alzheimer gaixotasunean eta gaixotasun honetako zenbait animalia-eredutan. Aldaketa horien zentzua oraindik ez dago finkatuta, baina haien eragina beste neurotransmisio-sistemen edota bestelako funtzio biologikoen modulazioaren bidez gerta daitezke. Beraz, lipido hauek etorkizun handiko itu farmakologikoak izan daitezke Alzheimer gaixotasunean agertzen diren sintoma neuropatologikoak eta neuropsikiatrikoak arintzeko. Hortaz, merkatuan dauden lipidoen seinaleztapena itutzat duten eta beste neuroendekapenezko gaixotasunak tratatzeko erabilgarriak diren farmakoak baliagarriak izan litezke Alzheimer gaixotasuna tratatzeko, aukera emanez horrela Alzheimer gaixotasuna tratatzeko dagoen hutsune farmakologikoa betetzeko.; In addition to energy and structural functions, lipids are becoming important thanks to the other functions described. Some lipids have been shown to exhibit neurotransmitter or neuromodulatory function, including lysophospholipids. Lysophospholipids are small bioactive lipid molecules that are distinguished only by having a single carbon chain and a single polar head. The lysophosphatidic acid and sphingosine l-phosphate structure and system functions are best described among those with neurotransmitter function. Lysophospholipids act as extracellular mediators that activate receptor-specific G proteins that are specific to them. The signaling of these molecules modulate certain neurochemical processes, including neuromodulation and neuroinflammation. They have also presented the relationship with learning and memory. In this respect, the best described lysophospholipid systems, lysophosphatidic acid and sphingosine 1-phosphate, are indeed disturbed in Alzheimer’s disease and in some animal models of this disease. The meaning of these changes is not yet established, but their effect may be related to the modulation of other neurotransmission systems or other biological functions. These lipids are therefore supposed to be the promising pharmacological targets to alleviate the neuropathological and neuropsychiatric symptoms that appear in Alzheimer’s disease. Therefore, marketed drugs that have lipid signaling as a pharmacological target and that are useful to treat other neurodegenerative diseases could be also helpful to treat the Alzheimer’s disease, and with this it might be possible to fill the pharmacological gap in the treatment of Alzheimer’s disease so far

Sexuality, gender, religion and interculturality in Spanish television civilising and cultural news stories

Introducción. En esta investigación se analizan distintas noticias sobre sexualidad, género, religión e interculturalidad en televisiones españolas. Metodología. Se realiza un análisis del discurso de las distintas identidades enmarcadas en las mencionadas categorías. A continuación, en distintos grupos discusión se analizan las interpretaciones que hacen las audiencias de dichas noticias. Finalmente, a su vez, analizamos mediante el método Delphi la opinión de distintos expertos sobre estas mismas noticias. Resultados y Discusión. Gracias al análisis del discurso se desvelan las representaciones y los estereotipos mediáticos que las audiencias y expertos identifican fácilmente. Conclusiones. Hay que señalar que tanto las audiencias como los expertos fueron muy críticos con las piezas sometidas a su interpretación, y percibieron claramente las insuficiencias y los estereotipos utilizados. Sin embargo, sobre todo las audiencias de mayor edad, a la hora de abordar dichos temas en general suelen tener los mismos prejuicios que denuncian en los informativos.Introduction. This paper analyzes several Spanish TV news about sexuality, gender, religion and interculturality to explore how these news are interpreted by audiences and experts. Methodology. We have used three complementary methods. First, we have conducted a critical discourse analysis (CDA) of different sexual, religious and intercultural identities portrayed by the TV news. We have also analyzed how audiences interpret these news through focus groups. Finally, we have used a delphi method to analyze the interpretation of TV experts. Results and Discussion. The CDA illustrated common stereotypes identified by audiences and experts. Conclusions. Audiences and experts were very critical with TV news and perceived the limitations and stereotypes portrayed. However, older audiences used the same stereotypes demonstrated in the focus groups. In general, this study shows that when receivers interpret sensitive issues, they trend to accept or reject media discourses, letting little space for the negotiation of meanings

Cannabinoid signaling modulation through JZL184 restores key phenotypes of a mouse model for Williams-Beuren syndrome

Williams-Beuren syndrome (WBS) is a rare genetic multisystemic disorder characterized by mild-to-moderate intellectual disability and hypersocial phenotype, while the most life-threatening features are cardiovascular abnormalities. Nowadays, there are no pharmacological treatments to directly ameliorate the main traits of WBS. The endocannabinoid system (ECS), given its relevance for both cognitive and cardiovascular function, could be a potential druggable target in this syndrome. We analyzed the components of the ECS in the complete deletion (CD) mouse model of WBS and assessed the impact of its pharmacological modulation in key phenotypes relevant for WBS. CD mice showed the characteristic hypersociable phenotype with no preference for social novelty and poor short-term object-recognition performance. Brain cannabinoid type-1 receptor (CB1R) in CD male mice showed alterations in density and coupling with no detectable change in main endocannabinoids. Endocannabinoid signaling modulation with subchronic (10 days) JZL184, a selective inhibitor of monoacylglycerol lipase, specifically normalized the social and cognitive phenotype of CD mice. Notably, JZL184 treatment improved cardiovascular function and restored gene expression patterns in cardiac tissue. These results reveal the modulation of the ECS as a promising novel therapeutic approach to improve key phenotypic alterations in WBS