Amyotrophic Lateral Sclerosis ALS and adenosine receptors

Copyright © 2018 Sebastião, Rei and Ribeiro. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.In the present review we discuss the potential involvement of adenosinergic signaling, in particular the role of adenosine receptors, in amyotrophic lateral sclerosis (ALS). Though the literature on this topic is not abundant, the information so far available on adenosine receptors in animal models of ALS highlights the interest to continue to explore the role of these receptors in this neurodegenerative disease. Indeed, all motor neurons affected in ALS are responsive to adenosine receptor ligands but interestingly, there are alterations in pre-symptomatic or early symptomatic stages that mirror those in advanced disease stages. Information starts to emerge pointing toward a beneficial role of A2A receptors (A2AR), most probably at early disease states, and a detrimental role of caffeine, in clear contrast with what occurs in other neurodegenerative diseases. However, some evidence also exists on a beneficial action of A2AR antagonists. It may happen that there are time windows where A2AR prove beneficial and others where their blockade is required. Furthermore, the same changes may not occur simultaneously at the different synapses. In line with this, it is not fully understood if ALS is a dying back disease or if it propagates in a centrifugal way. It thus seems crucial to understand how motor neuron dysfunction occurs, how adenosine receptors are involved in those dysfunctions and whether the early changes in purinergic signaling are compensatory or triggers for the disease. Getting this information is crucial before starting the design of purinergic based strategies to halt or delay disease progression.This work was supported by LISBOA-01-0145-FEDER-007391, project co-funded by FEDER through POR Lisboa 2020 (Programa Operacional Regional de Lisboa) from PORTUGAL 2020 and Fundação para a Ciência e Tecnologia (FCT) and by a Twinning action (SynaNet) from the EU H2020 program (Project Number: 692340). NR is in receipt of an FCT fellowship (PD /BD/113463/2015) and is a fellow of the M2B-Ph.D. Program.info:eu-repo/semantics/publishedVersio

Changes in adenosine receptors and neurotrophic factors in the SOD1G93A mouse model of amyotrophic lateral sclerosis: modulation by chronic caffeine

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of corticospinal tract motor neurons. Previous studies showed that adenosine-mediated neuromodulation is disturbed in ALS and that vascular endothelial growth factor (VEGF) has a neuroprotective function in ALS mouse models. We evaluated how adenosine (A1R and A2AR) and VEGF (VEGFA, VEGFB, VEGFR-1 and VEGFR-2) system markers are altered in the cortex and spinal cord of pre-symptomatic and symptomatic SOD1G93A mice. We then assessed if/how chronic treatment of SOD1G93A mice with a widely consumed adenosine receptor antagonist, caffeine, modulates VEGF system and/or the levels of Brain-derived Neurotrophic Factor (BDNF), known to be under control of A2AR. We found out decreases in A1R and increases in A2AR levels even before disease onset. Concerning the VEGF system, we detected increases of VEGFB and VEGFR-2 levels in the spinal cord at pre-symptomatic stage, which reverses at the symptomatic stage, and decreases of VEGFA levels in the cortex, in very late disease states. Chronic treatment with caffeine rescued cortical A1R levels in SOD1G93A mice, bringing them to control levels, while rendering VEGF signaling nearly unaffected. In contrast, BDNF levels were significantly affected in SOD1G93A mice treated with caffeine, being decreased in the cortex and increased in spinal the cord. Altogether, these findings suggest an early dysfunction of the adenosinergic system in ALS and highlights the possibility that the negative influence of caffeine previously reported in ALS animal models results from interference with BDNF rather than with the VEGF signaling molecules.info:eu-repo/semantics/publishedVersio

Unexpected short- and long-term effects of chronic adolescent HU-210 exposure on emotional behavior

© 2022 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by- nc-nd/4.0/).Chronic adolescent cannabinoid receptor agonist exposure has been shown to lead to persistent increases in depressive-like behaviors. This has been a key obstacle to the development of cannabinoid-based therapeutics. However, most of the published work has been performed with only three compounds, namely Δ9-tetrahydrocannabinol, CP55,940 and WIN55,212-2. Hypothesizing that different compounds may lead to distinct outcomes, we herein used the highly potent CB1R/CB2R full agonist HU-210, and first aimed at replicating cannabinoid-induced long-lasting effects, by exposing adolescent female Sprague-Dawley rats to increasing doses of HU-210, for 11 days and testing them at adulthood, after a 30-day drug washout. Surprisingly, HU-210 did not significantly impact adult anxious- or depressive-like behaviors. We then tested whether chronic adolescent HU-210 treatment resulted in short-term (24h) alterations in depressive-like behavior. Remarkably, HU-210 treatment simultaneously induced marked antidepressant- and prodepressant-like responses, in the modified forced swim (mFST) and sucrose preference tests (SPT), respectively. Hypothesizing that mFST results were a misleading artifact of HU-210-induced behavioral hyperreactivity to stress, we assessed plasmatic noradrenaline and corticosterone levels, under basal conditions and following an acute swim-stress episode. Notably, we found that while HU-210 did not alter basal noradrenaline or corticosterone levels, it greatly augmented the stress-induced increase in both. Our results show that, contrary to previously studied cannabinoid receptor agonists, HU-210 does not induce persisting depressive-like alterations, despite inducing marked short-term increases in stress-induced reactivity. By showing that not all cannabinoid receptor agonists may induce long-term negative effects, these results hold significant relevance for the development of cannabinoid-based therapeutics.Work was supported by project funding from Fundação para a Ciência e para a Tecnologia (FCT) (PTDC/MED-FAR/30933/2017 and PTDC/MED-FAR/4834/2021) and by H2020-WIDESPREAD-05-2017-Twinning (EpiEpinet) under grant agreement No. 952455. MF-F (SFRH/BD/147505/2019), NR (PD/BD/113463/2015), JF-G (PD/BD/114441/2016) and CM-L (SFRH/BD/118238/2016) are supported by PhD fellowships from FCT. The funding sources had no involvement in study design, preparation of the manuscript, or decision regarding its submission.info:eu-repo/semantics/publishedVersio

Going the extra (synaptic) mile: excitotoxicity as the road toward neurodegenerative diseases

Copyright © 2020 Armada-Moreira, Gomes, Pina, Savchak, Gonçalves-Ribeiro, Rei, Pinto, Morais, Martins, Ribeiro, Sebastião, Crunelli and Vaz. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.Excitotoxicity is a phenomenon that describes the toxic actions of excitatory neurotransmitters, primarily glutamate, where the exacerbated or prolonged activation of glutamate receptors starts a cascade of neurotoxicity that ultimately leads to the loss of neuronal function and cell death. In this process, the shift between normal physiological function and excitotoxicity is largely controlled by astrocytes since they can control the levels of glutamate on the synaptic cleft. This control is achieved through glutamate clearance from the synaptic cleft and its underlying recycling through the glutamate-glutamine cycle. The molecular mechanism that triggers excitotoxicity involves alterations in glutamate and calcium metabolism, dysfunction of glutamate transporters, and malfunction of glutamate receptors, particularly N-methyl-D-aspartic acid receptors (NMDAR). On the other hand, excitotoxicity can be regarded as a consequence of other cellular phenomena, such as mitochondrial dysfunction, physical neuronal damage, and oxidative stress. Regardless, it is known that the excessive activation of NMDAR results in the sustained influx of calcium into neurons and leads to several deleterious consequences, including mitochondrial dysfunction, reactive oxygen species (ROS) overproduction, impairment of calcium buffering, the release of pro-apoptotic factors, among others, that inevitably contribute to neuronal loss. A large body of evidence implicates NMDAR-mediated excitotoxicity as a central mechanism in the pathogenesis of many neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), and epilepsy. In this review article, we explore different causes and consequences of excitotoxicity, discuss the involvement of NMDAR-mediated excitotoxicity and its downstream effects on several neurodegenerative disorders, and identify possible strategies to study new aspects of these diseases that may lead to the discovery of new therapeutic approaches. With the understanding that excitotoxicity is a common denominator in neurodegenerative diseases and other disorders, a new perspective on therapy can be considered, where the targets are not specific symptoms, but the underlying cellular phenomena of the disease.This work was funded by Fundação para a Ciência e Tecnologia (FCT), Portugal (PTDC/BTM-SAL/32147/2017, PD/BD/114278/2016, IMM/BI/2-2020), and by FCT/Ministério da Ciência, Tecnologia e Ensino Superior (MCTES) through Fundos do Orçamento de Estado (UID/BIM/50005/2019). TM was funded by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska/Curie grant agreement No. 722053.info:eu-repo/semantics/publishedVersio

Characterization of the Neuromodulatory Dysfunction in Amyotrophic Lateral Sclerosis

Programa Doutoral M2B-PhD Medical Biochemistry and Biophysic

Diagnóstico molecular de formas familiares de hipercalcémia

Tese de mestrado, Biologia (Biologia Molecular e Genética), 2008, Universidade de Lisboa, Faculdade de CiênciasA hipersecreção de parathormona (PTH), associada a hipercalcémia, constitui uma endocrinopatia designada por hiperparatiroidismo primário (HPT), que pode resultar da hiperactividade de adenomas (80-85%), hiperplasia (15-20%) ou carcinomas (cerca de 1%) das paratiróides. O hiperparatiroidismo surge com maior frequência na forma esporádica, no entanto, cerca de 10% dos doentes apresentam uma forma familiar. Na presente tese, pretendeu-se identificar e caracterizar mutações germinais nos genes CaSR, HRPT2 e MEN1, envolvidos em formas familiares de hipercalcémia, respectivamente, hipercalcémia e hipocalciúria familiar (FHH)/hiperparatiroidismo neonatal severo (NSHPT), hiperparatiroidismo e tumores nos maxilares (HPT-JT) e neoplasias endócrinas múltiplas de tipo 1 (MEN1). Foram estudados dez casos índex e dezanove familiares destes mesmos casos. A sequenciação do gene CaSR, revelou duas novas mutações (Arg69His e c.1945delG) e uma mutação já descrita na literatura (Arg680His), em duas famílias e num caso sem história familiar, com FHH/NSHPT. Em dois casos com tumores das paratiróides, sem história familiar da doença, foram identificadas duas novas mutações germinais no gene HRPT2 (Arg76stop e c.518del4bp), sugerindo que as lesões apresentadas por estes indivíduos representavam uma expressão incompleta da síndrome HPT-JT. A detecção de uma mutação somática (Leu5Pro) no gene HRPT2, num carcinoma da paratiróide de uma doente portadora de uma mutação germinal neste gene, confirmou a inactivação total e a função oncosupressora do gene HRPT2. Estudos de segregação com marcadores de microssatélites, em dois novos indivíduos de uma família previamente estudada, em que tinha sido detectada uma extensa delecção germinal no gene MEN1, del (exão 7 região 3' não traduzida), revelaram que estes indivíduos não tinham herdado o haplotipo associado ao alelo mutado. O estudo desenvolvido neste trabalho permitiu não só a identificação precoce de dez portadores de mutações germinais em genes envolvidos na hipercalcémia familiar, mas também uma melhor compreensão dos mecanismos moleculares envolvidos na tumorigénese das paratiróidesThe hypersecretion of parathormone (PTH), associated to hypercalcemia, is an endocrinopathy designated as primary hyperparathyroidism (HPT), which may result from the hyperactivity of adenomas (80-85%), hyperplasia (15-20%) or carcinomas (about 1 %) of the parathyroid glands. Primary hyperparathyroidism usually occurs sporadically, however, approximately 10% of the patients present a familial form. The aim of the present study was to identify and characterize germ line mutations in the CaSR, HRPT2 and MEN1 genes, which are involved in familial forms of hypercalcemia, respectively, familial hypocalciuric hypercalcemia (FHH)/neonatal severe hyperparathyroidism (NSHPT), hyperparathyroidism-jaw tumour (HPT-JT), and multiple endocrine neoplasia type 1 (MEN1). Ten probands and nineteen relatives were studied. Sequencing analysis of the CaSR gene revealed two novel mutations (Arg69His and c.1945delG) and one previously described (Arg680His), in two families and one case with no family history, with FHH/NSHPT. In two cases presenting parathyroid tumours, with no family history of the disease, two novel germ line mutations in the HRPT2 gene (Arg76stop and c.518del4bp) were identified, suggesting that the parathyroid lesions represented an incomplete expression of HPT-JT syndrome. The detection of a somatic HRPT2 mutation (Leu5Pro) in the parathyroid carcinoma of a patient carrying a germ line mutation in this gene, confirmed the total inactivation and the oncosupression function of the HRPT2 gene. Segregation studies, using microsatellites markers, in two new individuals, from a previously studied family with a large deletion in the MEN1 gene, del (exon 7 3' non translated region), revealed that these individuals did not have inherited the haplotype associated to the mutant allele. The study developed in the present thesis, allowed the early identification of ten individuals carrying germ line mutations in genes involved in familial hypercalcemia and contributed to a better understanding of molecular mechanisms involved in parathyroid tumorigenesi

Amyotrophic Lateral Sclerosis (ALS) and Adenosine Receptors

In the present review we discuss the potential involvement of adenosinergic signaling, in particular the role of adenosine receptors, in amyotrophic lateral sclerosis (ALS). Though the literature on this topic is not abundant, the information so far available on adenosine receptors in animal models of ALS highlights the interest to continue to explore the role of these receptors in this neurodegenerative disease. Indeed, all motor neurons affected in ALS are responsive to adenosine receptor ligands but interestingly, there are alterations in pre-symptomatic or early symptomatic stages that mirror those in advanced disease stages. Information starts to emerge pointing toward a beneficial role of A2A receptors (A2AR), most probably at early disease states, and a detrimental role of caffeine, in clear contrast with what occurs in other neurodegenerative diseases. However, some evidence also exists on a beneficial action of A2AR antagonists. It may happen that there are time windows where A2AR prove beneficial and others where their blockade is required. Furthermore, the same changes may not occur simultaneously at the different synapses. In line with this, it is not fully understood if ALS is a dying back disease or if it propagates in a centrifugal way. It thus seems crucial to understand how motor neuron dysfunction occurs, how adenosine receptors are involved in those dysfunctions and whether the early changes in purinergic signaling are compensatory or triggers for the disease. Getting this information is crucial before starting the design of purinergic based strategies to halt or delay disease progression

Changes in adenosine receptors and neurotrophic factors in the SOD1G93A mouse model of amyotrophic lateral sclerosis: Modulation by chronic caffeine.

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of corticospinal tract motor neurons. Previous studies showed that adenosine-mediated neuromodulation is disturbed in ALS and that vascular endothelial growth factor (VEGF) has a neuroprotective function in ALS mouse models. We evaluated how adenosine (A1R and A2AR) and VEGF (VEGFA, VEGFB, VEGFR-1 and VEGFR-2) system markers are altered in the cortex and spinal cord of pre-symptomatic and symptomatic SOD1G93A mice. We then assessed if/how chronic treatment of SOD1G93A mice with a widely consumed adenosine receptor antagonist, caffeine, modulates VEGF system and/or the levels of Brain-derived Neurotrophic Factor (BDNF), known to be under control of A2AR. We found out decreases in A1R and increases in A2AR levels even before disease onset. Concerning the VEGF system, we detected increases of VEGFB and VEGFR-2 levels in the spinal cord at pre-symptomatic stage, which reverses at the symptomatic stage, and decreases of VEGFA levels in the cortex, in very late disease states. Chronic treatment with caffeine rescued cortical A1R levels in SOD1G93A mice, bringing them to control levels, while rendering VEGF signaling nearly unaffected. In contrast, BDNF levels were significantly affected in SOD1G93A mice treated with caffeine, being decreased in the cortex and increased in spinal the cord. Altogether, these findings suggest an early dysfunction of the adenosinergic system in ALS and highlights the possibility that the negative influence of caffeine previously reported in ALS animal models results from interference with BDNF rather than with the VEGF signaling molecules

A₁R and A_2AR mRNA expression and protein levels in the cortex of pre-symptomatic and symptomatic SOD1^G93A mice.

(A) Cortical A1R mRNA expression in pre-symptomatic (4–6 weeks old) and symptomatic (12–14 weeks old) SOD1G93A and age-matched WT mice. The Y-axis represents the A1R transcript level of the normalized A1R/GAPDH ratio in each condition. (B) Saturation binding curves of A1R for pre-symptomatic and symptomatic SOD1G93A and age-matched WT mice, in cortex. The Y-axis represents the specific binding of A1R expressed in fmol/mg protein plotted against increased concentrations of [3H] DPCPX (nM) (abscissae). (C) Maximum density of A1R (Bmax) in cortex of pre-symptomatic and symptomatic SOD1G93A and age-matched WT mice. (D) DPCPX equilibrium dissociation constant (KD) expressed in nM, for pre-symptomatic and symptomatic SOD1G93A and age-matched WT mice, in cortex. (E) Cortical A2AR mRNA expression in pre-symptomatic and symptomatic SOD1G93A and age-matched WT mice. The Y-axis represents the A2AR transcript level of the normalized A2AR/GAPDH ratio in each condition. (F) Cortical A2AR protein levels in pre-symptomatic and symptomatic SOD1G93A and age-matched WT mice. The Y-axis represents the A2AR/GAPDH ratios normalized to age-matched WT samples. Upper panels show representative immunoblots for each condition. Data are expressed as mean ±SEM (n = 4–7 animals for all conditions); *p < 0.05, **p ≤ 0.01, ***p ≤ 0.001, two-way ANOVA with Holm-Sidak correction for multiple comparisons.</p

Raw_Western blots images and analysis.

(PDF)</p