Academic literature on the topic 'Volume regulation; Neuronal cells; Brain cells'

The mammalian brain is composed of four distinct fluid compartments: blood, cerebral spinal fluid, interstitial fluid surrounding glial cells and neurons, and intracellular fluid. Maintenance of the ionic and osmotic composition and volume of these fluids is crucial for the normal functioning of the brain. Small changes in intracellular or extracellular solute composition can dramatically alter neuronal signaling and information processing. Because of the rigid confines of the skull and complex brain architecture, changes in total brain volume can cause devastating neurological damage. As a result, it is not surprising to find that the composition and volume of brain intracellular and extracellular fluids are controlled tightly under both normal conditions and in various disease states. Osmotic and ionic balance in the central nervous system is regulated by solute and water transport across the blood-brain barrier, the choroid plexus, and the plasma membrane of glial cells and neurons. Despite its clinical and physiological significance, however, little is known about the underlying cellular and molecular mechanisms by which the central nervous system's osmotic and ionic balance is maintained. In this review, the current understanding of osmoregulation in the mammalian brain and its role in various disease processes such as hyponatremia, renal failure, and hypernatremia will be summarized. A detailed understanding of brain osmoregulatory processes represents a fundamental physiological problem and is required for the treatment of numerous disease states, particularly those encountered in the practice of nephrology.

Na+-K+-2Cl−cotransporters are important in renal salt reabsorption and in salt secretion by epithelia. They are also essential in maintenance and regulation of ion gradients and cell volume in both epithelial and nonepithelial cells. Expression of Na+-K+-2Cl−cotransporters in brain tissues is high; however, little is known about their function and regulation in neurons. In this study, we examined regulation of the Na+-K+-2Cl−cotransporter by the excitatory neurotransmitter glutamate. The cotransporter activity in human neuroblastoma SH-SY5Y cells was assessed by bumetanide-sensitive K+ influx, and protein expression was evaluated by Western blot analysis. Glutamate was found to induce a dose- and time-dependent stimulation of Na+-K+-2Cl−cotransporter activity in SH-SY5Y cells. Moreover, both the glutamate ionotropic receptor agonist N-methyl-d-aspartic acid (NMDA) and the metabotropic receptor agonist (±)-1-aminocyclopentane- trans-1,3-dicarboxylic acid ( trans-ACPD) significantly stimulated the cotransport activity in these cells. NMDA-mediated stimulation of the Na+-K+-2Cl−cotransporter was abolished by the selective NMDA-receptor antagonist (+)-MK-801 hydrogen maleate. trans-ACPD-mediated effect on the cotransporter was blocked by the metabotropic receptor antagonist (+)-α-methyl-(4-carboxyphenyl)glycine. The results demonstrate that Na+-K+-2Cl−cotransporters in neurons are regulated by activation of both ionotropic and metabotropic glutamate receptors.

Hyponatremia is the most common electrolyte abnormality in hospitalized patients. When symptomatic (hyponatremic encephalopathy), the overall morbidity is 34%. Individuals most susceptible to death or permanent brain damage are prepubescent children and menstruant women. Failure of the brain to adapt to the hyponatremia leads to brain damage. Major factors that can impair brain adaptation include hypoxia and peptide hormones. In children, physical factors—discrepancy between skull size and brain size—are important in the genesis of brain damage. In adults, certain hormones—estrogen and vasopressin (usually elevated in cases of hyponatremia)—have been shown to impair brain adaptation, decreasing both cerebral blood flow and oxygen utilization. Initially, hyponatremia leads to an influx of water into the brain, primarily through glial cells and largely via the water channel aquaporin (AQP)4. Water is thus shunted into astrocytes, which swell, largely preserving neuronal cell volume. The initial brain response to swelling is adaptation, utilizing the Na+-K+-ATPase system to extrude cellular Na+. In menstruant women, estrogen + vasopressin inhibits the Na+-K+-ATPase system and decreases cerebral oxygen utilization, impairing brain adaptation. Cerebral edema compresses the respiratory centers and also forces blood out of the brain, both lowering arterial Po2 and decreasing oxygen utilization. The hypoxemia further impairs brain adaptation. Hyponatremic encephalopathy leads to brain damage when brain adaptation is impaired and is a consequence of both cerebral hypoxia and peptide hormones.

AbstractMany diverse retinal disorders are characterized by retinal edema; yet, little experimental attention has been given to understanding the fundamental mechanisms underlying and contributing to these fluid-based disorders. Water transport in and out of cells is achieved by specialized membrane channels, with most rapid water transport regulated by transmembrane water channels known as aquaporins (AQPs). The predominant AQP in the mammalian retina is AQP4, which is expressed on the Müller glial cells. Müller cells have previously been shown to modulate neuronal activity by modifying the concentrations of ions, neurotransmitters, and other neuroactive substances within the extracellular space between the inner and the outer limiting membrane. In doing so, Müller cells maintain extracellular homeostasis, especially with regard to the spatial buffering of extracellular potassium (K+) via inward rectifying K+ channels (Kir channels). Recent studies of water transport and the spatial buffering of K+ through glial cells have highlighted the involvement of both AQP4 and Kir channels in regulating the extracellular environment in the brain and retina. As both glial functions are associated with neuronal activation, controversy exists in the literature as to whether the relationship is functionally dependent. It is argued in this review that as AQP4 channels are likely to be the conduit for facilitating fluid homeostasis in the inner retina during light activation, AQP4 channels are also likely to play a consequent role in the regulation of ocular volume and growth. Recent research has already shown that the level of AQP4 expression is associated with environmentally driven manipulations of light activity on the retina and the development of myopia.

Potassium is tightly regulated within the extracellular compartment of the brain. Nonetheless, it can increase 3- to 4-fold during periods of intense seizure activity and 10- to 20-fold under certain pathological conditions such as spreading depression. Within the central nervous system, neurons and astrocytes are both affected by shifts in the extracellular concentration of potassium. Elevated potassium can lead to a redistribution of other ions (e.g., calcium, sodium, chloride, hydrogen, etc.) within the cellular compartment of the brain. Small shifts in the extracellular potassium concentration can markedly affect acid–base homeostasis, energy metabolism, and volume regulation of these two brain cells. Since normal neuronal function is tightly coupled to the ability of the surrounding glial cells to regulate ionic shifts within the brain and since both cell types can be affected by shifts in the extracellular potassium, it is important to characterize their individual response to an elevation of this ion. This review describes the results of side-by-side studies conducted on cortical neurons and astrocytes, which assessed the effect of elevated potassium on their resting membrane potential, intracellular volume, and their intracellular concentration of potassium, sodium, and chloride. The results obtained from these studies suggest that there exists a marked cellular heterogeneity between neurons and astrocytes in their response to an elevation in the extracellular potassium concentration.Key words: astrocytes, neurons, ion concentration, neuronal–glial interactions, mouse, cell culture.

AbstractThe small GTPase Rheb was originally detected as an immediate early response protein whose expression was induced by NMDA-dependent synaptic activity in the brain. Rheb’s activity is highly regulated by its GTPase activating protein (GAP), the tuberous sclerosis complex protein, which stimulates the conversion from the active, GTP-loaded into the inactive, GDP-loaded conformation. Rheb has been established as an evolutionarily conserved molecular switch protein regulating cellular growth, cell volume, cell cycle, autophagy, and amino acid uptake. The subcellular localization of Rheb and its interacting proteins critically regulate its activity and function. In stem cells, constitutive activation of Rheb enhances differentiation at the expense of self-renewal partially explaining the adverse effects of deregulated Rheb in the mammalian brain. In the context of various cellular stress conditions such as oxidative stress, ER-stress, death factor signaling, and cellular aging, Rheb activation surprisingly enhances rather than prevents cellular degeneration. This review addresses cell type- and cell state-specific function(s) of Rheb and mainly focuses on neurons and their surrounding glial cells. Mechanisms will be discussed in the context of therapy that interferes with Rheb’s activity using the antibiotic rapamycin or low molecular weight compounds.

Brain injury is a significant risk factor for chronic gliosis and neurodegenerative diseases. Currently, no treatment is available for neuroinflammation caused by the action of glial cells following brain injury. In this study, we investigated the quinpirole-mediated activation of dopamine D2 receptors (D2R) in a mouse model of traumatic brain injury (TBI). We also investigated the neuroprotective effects of quinpirole (a D2R agonist) against glial cell-induced neuroinflammation secondary to TBI in adult mice. After the brain injury, we injected quinpirole into the TBI mice at a dose of 1 mg/kg daily intraperitoneally for 7 days. Our results showed suppression of D2R expression and deregulation of downstream signaling molecules in ipsilateral cortex and striatum after TBI on day 7. Quinpirole administration regulated D2R expression and significantly reduced glial cell-induced neuroinflammation via the D2R/Akt/glycogen synthase kinase 3 beta (GSK3-β) signaling pathway after TBI. Quinpirole treatment concomitantly attenuated increase in glial cells, neuronal apoptosis, synaptic dysfunction, and regulated proteins associated with the blood–brain barrier, together with the recovery of lesion volume in the TBI mouse model. Additionally, our in vitro results confirmed that quinpirole reversed the microglial condition media complex-mediated deleterious effects and regulated D2R levels in HT22 cells. This study showed that quinpirole administration after TBI reduced secondary brain injury-induced glial cell activation and neuroinflammation via regulation of the D2R/Akt/GSK3-β signaling pathways. Our study suggests that quinpirole may be a safe therapeutic agent against TBI-induced neurodegeneration.

Portal hypertension due to either prehepatic portal hypertension or cirrhosis is associated with cardiovascular derangement. We aimed to delineate regulatory mechanisms in the brain stem cardiovascular nuclei in rat models of prehepatic portal hypertension and cirrhosis. Neuronal activation in the nucleus of the solitary tract (NTS) and ventrolateral medulla (VLM) were assessed by immunohistochemical staining for the immediate-early gene product Fos. In the same sections, catecholaminergic neurons were counted by tyrosine hydroxylase (TH) staining. Ninety minutes after hypotensive hemorrhage (or no volume challenge), the animals were killed for Fos and TH medullary staining. These protocols were repeated after capsaicin administration. The NTS of unchallenged sham-operated rats had scant Fos-positive cells (3.6 ± 0.4 cells/section), whereas hemorrhage significantly increased Fos staining (91.8 ± 14). In contrast, the unchallenged portal hypertensive and cirrhotic groups showed increased Fos staining (14.3 ± 5.8 and 32.8 ± 2.8, respectively), which hemorrhage did not alter significantly. The numbers of TH-positive cells were similar in the three unchallenged groups; double labeling revealed that ∼50% of TH-positive cells were activated by hemorrhage in the sham and cirrhotic rats but not the portal hypertensive rats. Similar patterns of Fos and TH staining were observed in the VLM. Capsaicin treatment not only significantly reduced the Fos-positive neuron numbers in portal hypertensive and cirrhotic rats but also attenuated hemorrhage-induced Fos and double-positive cells in both NTS and VLM. These results suggest that disordered trafficking in capsaicin-sensitive nerves and central dysregulation contribute to blunted cardiovascular responsiveness in cirrhosis and prehepatic portal hypertension.

The pursuit of targeting multiple pathways in the ischemic cascade of cerebral stroke is a promising treatment option. We examined the regenerative potential of conditioned medium derived from rat and human apoptotic mononuclear cells (MNC), rMNCapo sec and hMNCapo sec, in experimental stroke.We performed middle cerebral artery occlusion on Wistar rats and administered apoptotic MNC-secretomes intraperitoneally in two experimental settings. Ischemic lesion volumes were determined 48 hours after cerebral ischemia. Neurological evaluations were performed after 6, 24 and 48 hours. Immunoblots were conducted to analyze neuroprotective signal-transduction in human primary glia cells and neurons. Neuronal sprouting assays were performed and neurotrophic factors in both hMNCapo sec and rat plasma were quantified using ELISA.Administration of rat as well as human apoptotic MNC-secretomes significantly reduced ischemic lesion volumes by 36% and 37%, respectively. Neurological examinations revealed improvement after stroke in both treatment groups. Co-incubation of human astrocytes, Schwann cells and neurons with hMNCapo sec resulted in activation of several signaling cascades associated with the regulation of cytoprotective gene products and enhanced neuronal sprouting in vitro. Analysis of neurotrophic factors in hMNCapo sec and rat plasma revealed high levels of brain derived neurotrophic factor (BDNF).Our data indicate that apoptotic MNC-secretomes elicit neuroprotective effects on rats that have undergone ischemic stroke.

Astrocytes and neurons have been shown to swell across a variety of different conditions, including increases in extracellular potassium concentration (^[K+]o). The mechanisms involved in the coupling of K+ influx to water movement into cells leading to cell swelling are not well understood and remain controversial. Here, we set out to determine the effects of ^[K+]o on rapid volume responses of hippocampal CA1 pyramidal neurons and stratum radiatum astrocytes using real-time confocal volume imaging. First, we found that elevating [K+]o within a physiological range (to 6.5 mM and 10.5 mM from a baseline of 2.5 mM), and even up to pathological levels (26 mM), produced dose-dependent increases in astrocyte volume, with absolutely no effect on neuronal volume. In the absence of compensating for addition of KCl by removal of an equal amount of NaCl, neurons actually shrank in ^[K+]o, while astrocytes continued to exhibit rapid volume increases. Astrocyte swelling in ^[K+]o was not dependent on neuronal firing, aquaporin 4, the inwardly rectifying potassium channel Kir 4.1, the sodium bicarbonate cotransporter NBCe1, , or the electroneutral cotransporter, sodium-potassium-chloride cotransporter type 1 (NKCC1), but was significantly attenuated in 1 mM barium chloride (BaCl2) and by the Na+/K+ ATPase inhibitor ouabain. Effects of 1 mM BaCl2 and ouabain applied together were not additive and, together with reports that BaCl2 can inhibit the NKA at high concentrations, suggests a prominent role for the astrocyte NKA in rapid astrocyte volume increases occurring in ^[K+]o. These findings carry important implications for understanding mechanisms of cellular edema, regulation of the brain extracellular space, and brain tissue excitability.

Introduction: The hBCAT proteins have a unique redox active CXXC motif which governs its transamination activity. Recent studies have identified that specific neuronal proteins with either redox activity or functions in cell signalling form interactions with the hBCAT proteins that are disrupted when the environment becoming oxidising. However, how hBCAT functions as an oxidoreductase is unknown or how it compares to other known cellular repair enzymes such as thioredoxin, glutaredoxin or protein disulphide isomerase. Moreover, leucine, a substrate for hBCAT regulates protein synthesis through the mTOR pathway, yet the importance of hBCAT itself in this mechanism remains undetermined. Aims: This thesis firstly investigated the redox substrates for the hBCAT proteins, and their oxidoreductase activity in comparison to the cellular repair enzymes. The second main aim was to establish if hBCAT can be regulated through phosphorylation both in-vitro and in the neuronal cell line (IMR-32). Finally, in addition to understanding their role as oxidoreductases, the importance of the reactive cysteines in redox binding to neuronal proteins was determined. Results: These studies demonstrated that both hBCAT isoforms have oxidoreductase and isomerase activity, but of lower activity relative to protein disulphide isomerase. The oxidoreductase activity was dependent on a functional CXXC motif, where in particular S-glutathionylation enhanced the ability of hBCAT to catalyse the folding of proteins. It has been demonstrated that hBCATc is regulated through phosphorylation and this is dependent on the redox environment unlike hBCATm which was only affected in the presence of thiol blocker, NEM. Both isoforms required the CXXC motif to be phosphorylated. Although phosphorylation of hBCATc in neuronal cells was observed, the exact mechanism needs to be elucidated. Finally, these studies have identified putative new partners for hBCAT such as inositol polyphosphate multikinase, GRINL1a upstream protein and parvalbumin. These indicate potential new roles for these proteins in cell division, neurotransmitter signal transduction and mTOR pathway, which have previously not been reported. Conclusion: The hBCAT proteins have oxidoreductase activity and are involved in a number of metabolic pathways such as cell division, neurotransmitter signal transduction and the mTOR pathway. This implies that these proteins may have a protective role in the cell, similarly to previous studies which have shown that upregulation of the hBCAT proteins is protective. Future work will elucidate further the role these proteins have under normal and pathological conditions.

Tese de doutoramento, Ciências Biomédicas (Neurociências), Universidade de Lisboa, Faculdade de Medicina, 2011
Gamma-aminobutyric acid (GABA) is the predominant inhibitory neurotransmitter in the central nervous system. Its activity at the synapse is terminated by re-uptake into nerve terminals and astrocytes, through membrane located specific GABA transporters (GATs), which therefore shape GABAergic transmission. There are three main high affinity subtypes of GATs, GAT-1, GAT-2 and GAT-3, and a low affinity one, the betaine transporter. GAT-1 is the predominant GABA transporter in the brain and is expressed in neurons and astrocytes. Several factors can regulate the continuous traffic of GATs to and from the neuronal plasma membrane. For instance surface expression of GAT-1 in cultured neurons and isolated nerve terminals is decreased by protein kinase C (PKC)-dependent phosphorylation. In contrast, surface expression of GAT-1 in neurons is enhanced by brain derived neurotrophic factor (BDNF)-mediated tyrosine kinase-dependent phosphorylation. Though reuptake of GABA might occur at different places of the neuronal membrane, its reuptake by the nerve terminal is the process that allows quick refilling of the released stores. On the other hand, uptake by astrocytes contributes to a fast removal of GABA from the synapse and delays its delivery to the neuronal release stores. To understand how GABAergic transmission can be shaped, it is therefore important to know how a single modulator can affect both processes of GABA removal from synapse. Thus, in the work presented on this thesis I evaluated the influence of BDNF upon GAT-1 transporters on presynaptic nerve terminals and cortical primary astrocyte culture. BDNF decreased GAT-1 mediated GABA uptake by isolated hippocampal rat nerve terminals (synaptosomes), an effect that occurred within 1 min of incubation with BDNF through activation of TrkB receptor. In contrast with what has been observed for other synaptic actions of BDNF, the inhibition of GABA transport by BDNF does not require tonic activation of adenosine A2A receptors, nevertheless is facilitated by activation of A2A receptors. On the other hand, BDNF enhances GAT-1 mediated GABA transport in cultured astrocytes, an effect mostly due to an increase in Vmax kinetic constant. This effect involves the truncated form of TrkB receptors (TrkB-t) coupled to a non-classic PLC-y/PKC-δ and Erk/MAP kinase pathway and requires active adenosine A2A receptors. To elucidate the trafficking of GAT-1 when astrocytes were treated with BDNF, a functional mutant of the rat GAT-1 was generated in which hemagglutinin epitope (HA) was incorporated into the second extracellular loop. By ELISA experiments, performed with astrocytes expressing HA-rGAT-1 transporter, it was possible to observe an exocytosis of HA-GAT-1 to plasma membrane when cells were treated with BDNF. In addition, cell surface biotinylation experiments, performed with astrocytes overexpressing the wild type rat GAT-1 (rGAT-1), also demonstrate an increase of GAT-1 2 transporter at plasma membrane when astrocytes were treated with BDNF. Results from experiments using selective inhibitors of endocytosis or selective inhibitors of recycling of molecules back to the plasma membrane allowed concluding that BDNF enhances GAT-1 expression at surface astrocytic membrane by slowing down exocytosis. A new role for BDNF is proposed whereby the effect of BDNF on GAT-1 transporter differs between pre-synaptic nerve terminals and astrocytes, suggesting that this neurotrophin operates in a much localized way, so that it may retard GABA uptake by the nerve terminal, enhancing synaptic actions of GABA, and accelerate its reuptake at extracellular neuronal areas allowing replenishment of neuronal pools of GABA. The results suggest that BDNF plays an active role in the regulation of GABAergic synaptic signalling, contributing to information processing.
O ácido y-aminobutírico (GABA) é o principal neurotransmissor inibitório do sistema nervoso central. A rápida remoção do GABA presente na fenda sináptica, por transportadores de alta afinidade para o GABA, que se localizam quer a nível do terminal pré-sináptico dos neurónios, quer a nível das células da glia, nomeadamenrte dos astrócitos (Gether et al., 2006), é essencial para uma sinalização eficaz mediada por este neurotransmissor. Até ao momento quatro transportadores foram identificados para o GABA, três destes de alta afinidade, denominados de GAT-1, GAT-2 and GAT-3 e um quarto, de baixa afinidade, denominado betaine transporter. O transportador GAT-1 é o transportador de GABA predominante no sistema nervoso central e encontra-se expresso preferencialmente em neurónios, sendo, no entanto, também expresso em astrócitos. Relativamente ao transportador GAT-3, sabe-se que este é maioritariamente expresso em astrócitos, onde tem um predomínio de transporte de GABA relativamente ao transportador GAT-1. Assim a recaptação de GABA pode ocorrer em diferentes localizações celulares. Quando ocorre para o terminal nervoso pre-sináptico, tem como consequência uma rápida reposição do nível de GABA nas vesículas sinápticas. A ocorrência para os astrócitos contribui para uma remoção mais rápida do GABA da fenda sináptica, diminuindo assim a velocidade de reposição de GABA nas vesículas sinápticas. Para se entender como é que a transmissão GABAérgica é regulada, torna-se pois extremamente relevante compreeender como pode apenas uma molécula modular os dois locais onde ocorre transporte de GABA, nomeadamente o pré-sináptico e o astrocítico. Salienta-se também a importância dos transportadores para o controlo da excitabilidade e o seu eventual envolvimento em situação patológica, nomeadamente em doentes com epilepsia do lobo temporal que apresentam uma aumento da expressão dos transportadores de GABA nos astrócitos. Os transportadores de GABA são regulados de diversos modos, estando envolvidos diferentes factores e várias cascatas de transdução de sinal. Esta modulação pode ocorrer de dois modos distintos: por alteração do Km ou da Vmax do transportador. A regulação do tráfego dos transportadores de GABA de, e para a membrana plasmática neuronal, pode ocorrer por variações da velocidade de endocitose e exocitose e/ou por alteração da quantidade de transportadores disponíveis neste processo de tráfego contínuo. Uma molécula já identificada como reguladora do transportador GAT-1 é o Brain derived neurotrophic factor (BDNF). O BDNF é um factor neurotrófico com importantes funções na diferenciação, maturação e sobrevivência neuronal, levando a modificações estruturais e moleculares a longo-prazo que são cruciais para o desenvolvimento, mas também para a função 2 e plasticidade sináptica no indivíduo adulto (Vicario-Abejon et al., 2002). O BDNF exerce a sua acção através da activação de receptores tirosina cinase B (TrkB), que se apresentam em diferentes isoformas: uma isoforma “completa” (TrkB-fl) que apresenta domínios tirosina cinase e uma isoforma truncada (TrkB-t) que não apresenta estes domínios. O BDNF favorece a recaptação de GABA devido a um aumento da expressão de GAT-1 a nível da membrana plasmática em culturas primárias de neurónios, não se sabendo até ao início deste trabalho qual a função do BDNF no controlo da actividade do GAT-1 local a nível de terminais nervosos. Os astrócitos são a maior classe de células da glia encontrada no cérebro dos mamíferos e têm um papel extremamente relevante na transmissão sináptica, contribuindo para o processamento de informação a nível sináptico ao controlar quer a composição do meio extracelular, quer a quantidade de neurotransmissores presentes na fenda sináptica. Os astrócitos são assim células fundamentais a nível da comunicação existente entre astrócitos ou entre astrócitos-neurónios. No que diz respeito à regulação dos níveis extracelulares de GABA, estas células têm um papel muito importante uma vez que expressam transportadores específicos de GABA, que permitem, como foi anteriormente referido, o controlo dos níveis deste neurotransmissor na fenda sináptica. Todavia, pouco tem sido descrito em relação à regulação dos transportadores de GABA nos astrócitos. O trabalho que aqui se apresenta teve como objectivo estudar o efeito do BDNF sobre o transportador de GABA, em terminais nervosos pré-sinápticos e em astrócitos, bem como estudar os mecanismos subjacentes ao efeito do BDNF. Foi também abordado o possível envolvimento dos receptores A2A da adenosina, uma vez que a interacção entre o receptor do BDNF, TrkB e o receptor de adenosina A2A, tem sido descrita em vários sistemas biológicos. Verificou-se que em terminais nervosos pré-sinápticos o BDNF tem uma acção inibitória sobre o transportador exclusivo de GABA (GAT-1) nesta estrutura, levando a uma diminuição da recaptação de GABA através deste transportador. Este efeito depende da concentração de BDNF e ocorre num intervalo de tempo extremamente curto (1 minuto). O efeito do BDNF no transportador GAT-1 ocorre através da activação do receptor TrkB e, contrariamente a outros efeitos mediados pela activação deste receptor, não requer a activação tónica dos receptores A2A da adenosina. Em culturas primárias de astrócitos o BDNF aumentou a recaptação de GABA mediada pelo transportador GAT-1, não tendo qualquer efeito no transportador GAT-3, também presente nos astrócitos. Este efeito ocorre devido a um aumento da velocidade máxima do transportador. O efeito do BDNF envolve a forma truncada do receptor TrkB, estando esta acoplada a uma via não clássica da PLC-y/PKC-δ e da Erk/MAP cinases. O efeito descrito requer que os receptores 3 A2A da adenosina estejam activos, sendo que os níveis endógenos de adenosina extracelular são suficientes para desencadear o efeito do BDNF. Uma vez que um aumento do Vmax se correlaciona com um aumento do número de transportadores na membrana plasmática, procedeu-se seguidamente à avaliação de um possível aumento da expressão do transportador GAT-1 quando as células eram tratadas com BDNF. Para avaliar se o efeito do BDNF se correlacionava com o tráfego de GAT-1 de, e para a membrana celular, foi gerado um mutante funcional do transportador GAT-1 de rato (rGAT-1), no qual foi introduzido o epítopo hemaglutinina (HA) no segundo loop extracelular do transportador, procedendo-se à infecção dos astrócitos com o referido mutante. Após o tratamento das células com BDNF observou-se um aumento da expressão de HA-rGAT-1 na membrana plasmática. Também através de experiências de biotinilação, realizadas com astrócitos que sobreexpressavam rGAT-1, se pôde concluir que o BDNF aumenta a expressão de rGAT-1 na membrana plasmática. Estudos onde se usou um inibidor da endocitose (dynasore) ou um inibidor da reciclagem de moléculas internalizadas de volta para a membrana plasmática (monensin), permitiram concluir que o efeito do BDNF envolve inibição da internalização de GAT-1 nos astrócitos, tendo esta acção consequências na expressão do GAT-1 e na velocidade de transporte de GABA. Os resultados apresentados nesta tese mostram que o BDNF exerce a sua acção de um modo muito localizado, levando a uma diminuição da recaptação de GABA no terminal nervoso que favorece eventualmente as suas acções sinápticas, e a uma aceleração da recaptação de GABA em regiões extra-sinápticas, que contribui para uma redução da acção tónica deste neurotransmissor. Em última instância, este efeito do BDNF deverá determinar uma diminuição da velocidade de reposição de GABA nas vesículas sinápticas, conduzindo desta forma a um aumento da excitabilidade neuronal.
Fundação para a Ciência e a Tecnologia (FCT, SFRH/BD/27989/2006)

No
The ability of neurons to modulate synaptic strength underpins synaptic plasticity, learning and memory, and adaptation to sensory experience. Despite the importance of synaptic adaptation in directing, reinforcing, and revising the behavioral response to environmental influences, the cellular and molecular mechanisms underlying synaptic adaptation are far from clear. Brain-derived neurotrophic factor (BDNF) is a prime initiator of structural and functional synaptic adaptation. However, the signaling cascade activated by BDNF to initiate these adaptive changes has not been elucidated. We have previously shown that BDNF activates mitogen- and stress-activated kinase 1 (MSK1), which regulates gene transcription via the phosphorylation of both CREB and histone H3. Using mice with a kinase-dead knock-in mutation of MSK1, we now show that MSK1 is necessary for the upregulation of synaptic strength in response to environmental enrichment in vivo. Furthermore, neurons from MSK1 kinase-dead mice failed to show scaling of synaptic transmission in response to activity deprivation in vitro, a deficit that could be rescued by reintroduction of wild-type MSK1. We also show that MSK1 forms part of a BDNF- and MAPK-dependent signaling cascade required for homeostatic synaptic scaling, which likely resides in the ability of MSK1 to regulate cell surface GluA1 expression via the induction of Arc/Arg3.1. These results demonstrate that MSK1 is an integral part of a signaling pathway that underlies the adaptive response to synaptic and environmental experience. MSK1 may thus act as a key homeostat in the activity- and experience-dependent regulation of synaptic strength.

Endocrine abnormalities are very common in patients with chronic autoimmune rheumatic diseases (CARDs) due to the systemic involvement of the central nervous system and endocrine glands. In recent years, the response of the endocrine (and also neuronal) system to peripheral inflammation has been linked to overall energy regulation of the diseased body and bioenergetics of immune cells. In CARDs, hormonal and neuronal pathways are outstandingly important in partitioning energy-rich fuels from muscle, brain, and fat tissue to the activated immune system. Neuroendocrine regulation of fuel allocation has been positively selected as an adaptive programme for transient serious, albeit non-life-threatening, inflammatory episodes. In CARDs, mistakenly, the adaptive programmes are used again but for a much longer time leading to systemic disease sequelae with endocrine (and also neuronal) abnormalities. The major endocrine alterations are depicted in the following list: mild activation of the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system, inadequate secretion of ACTH and cortisol relative to inflammation, loss of androgens, inhibition of the hypothalamic-pituitary-gonadal axis and fertility problems, high serum levels of oestrogens relative to androgens, fat deposits adjacent to inflamed tissue, increase of serum prolactin, and hyperinsulinaemia (and the metabolic syndrome). Neuroendocrine abnormalities are demonstrated using this framework that can explain many CARD-related endocrine disturbances. This chapter gives an overview on pathophysiology of neuroendocrine alterations in the context of energy regulation.

Endocrine abnormalities are very common in patients with chronic autoimmune rheumatic diseases (CARDs) due to the systemic involvement of the central nervous system and endocrine glands. In recent years, the response of the endocrine (and also neuronal) system to peripheral inflammation has been linked to overall energy regulation of the diseased body and bioenergetics of immune cells. In CARDs, hormonal and neuronal pathways are outstandingly important in partitioning energy-rich fuels from muscle, brain, and fat tissue to the activated immune system. Neuroendocrine regulation of fuel allocation has been positively selected as an adaptive programme for transient serious, albeit non-life-threatening, inflammatory episodes. In CARDs, mistakenly, the adaptive programmes are used again but for a much longer time leading to systemic disease sequelae with endocrine (and also neuronal) abnormalities. The major endocrine alterations are depicted in the following list: mild activation of the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system, inadequate secretion of ACTH and cortisol relative to inflammation, loss of androgens, inhibition of the hypothalamic-pituitary-gonadal axis and fertility problems, high serum levels of oestrogens relative to androgens, fat deposits adjacent to inflamed tissue, increase of serum prolactin, and hyperinsulinaemia (and the metabolic syndrome). Neuroendocrine abnormalities are demonstrated using this framework that can explain many CARD-related endocrine disturbances. This chapter gives an overview on pathophysiology of neuroendocrine alterations in the context of energy regulation.

Endocrine abnormalities are very common in patients with chronic autoimmune rheumatic diseases (CARDs) due to the systemic involvement of the central nervous system and endocrine glands. In recent years, the response of the endocrine (and also neuronal) system to peripheral inflammation has been linked to overall energy regulation of the diseased body and bioenergetics of immune cells. In CARDs, hormonal and neuronal pathways are outstandingly important in partitioning energy-rich fuels from muscle, brain, and fat tissue to the activated immune system. Neuroendocrine regulation of fuel allocation has been positively selected as an adaptive programme for transient serious, albeit non-life-threatening, inflammatory episodes. In CARDs, mistakenly, the adaptive programmes are used again but for a much longer time leading to systemic disease sequelae with endocrine (and also neuronal) abnormalities. The major endocrine alterations are depicted in the following list: mild activation of the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system, inadequate secretion of ACTH and cortisol relative to inflammation, loss of androgens, inhibition of the hypothalamic-pituitary-gonadal axis and fertility problems, high serum levels of oestrogens relative to androgens, fat deposits adjacent to inflamed tissue, increase of serum prolactin, and hyperinsulinaemia (and the metabolic syndrome). Neuroendocrine abnormalities are demonstrated using this framework that can explain many CARD-related endocrine disturbances. This chapter gives an overview on pathophysiology of neuroendocrine alterations in the context of energy regulation.

lSodium is the most abundant cation in the extracellular fluid and is important for regulation of plasma water concentrations and cell volume. Sodium cannot readily cross the blood-brain barrier, and changes in plasma sodium levels by altering free water movement can expand or shrink brain cells. Changes in brain cell volume can cause brain cell dysfunction and apoptosis. Correction of both high and low sodium levels must be done gradually, as rapid correction of dysnatremias can also damage brain cells. In this chapter we review the physiology of sodium regulation, and discuss the clinical implications of these disorders as well as present a treatment plan for safe correction.

Complex animals have evolved two separate systems for the control of body tissues. One is the nervous system, which makes direct connections with specific muscles and glands and regulates their activity by the focal release of neurotransmitters. The other system is the endocrine system, where hormones, secreted into the circulation, can exert effects on remote tissues in many different locations simultaneously. The classical distinction between the two systems is, however, blurred. Some hormones, such as antidiuretic hormone and oxytocin, are released into the bloodstream by neurones, rather than by typical endocrine cells. In other situations, hormones are released only to act locally, not all over the body, as with paracrine cells. Occasionally, the hormone feeds back on to the cell that secreted it, as in autocrine regulation. The interface between neural and endocrine control lies in the hypothalamus and related areas of the brain. This region also helps integrate the output of the autonomic nervous system, which controls visceral function. Hypothalamic areas also regulate appetite behaviours for food, water, sex, etc. Autonomic nervous system, appetites, and hormones all contribute to homeostasis — the regulation of the internal environment of the body. The hypothalamus and the pituitary gland form the ‘hypothalamic–pituitary endocrine axis’. This axis regulates much of the body’s endocrine activity through a system of hypothalamic factors. These factors, which are hormones in their own right, regulate the release of individual pituitary hormones. Each pituitary ‘trophic’ hormone then controls a part of the overall endocrine system. Thus, pituitary hormones control hormone production by thyroid, adrenal cortex, liver, and gonads. This complex cascade of hormonal control is regulated by various types of negative feedback based on plasma hormone concentrations. The hypothalamus and pituitary are also controlled by higher centres in the brain. Other endocrine tissues also use negative feedback control, but rather than the level of the hormone itself, it is the level of stimulus that regulates hormone secretion. Thus, rising plasma osmolarity (or decreasing blood volume) stimulates antidiuretic hormone secretion, and rising plasma glucose stimulates insulin secretion. Combinations of hormones are sometimes used to regulate an aspect of the internal environment. The control of plasma calcium by calcitonin, parathormone, and calcitriol (1,25-dihydroxycholecalciferol), and of plasma glucose by insulin and glucagon, are examples.

Neurodegeneration is the progressive and gradual dysfunction and loss of axons in the central nervous system. It is the main pathological characteristic of chronic and acute neurodegenerative conditions like Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS). The usual aspects of pathogenesis of disease can be abridged with regards to the downstream implications of uncontrollable protein oligomerization and aggregation from postmitotic cells. The brain structure constantly changes in normal aging without any dysfunction accompanying the structural changes in brain. The decline in cognitive capabilities, for example, processing speed, memory, and functions related to decision making are the sign of healthy aging. The reduction in brain volume in healthy aging is possibly related to neuronal loss at some marginal extent. The following chapter discusses the structural and functional alterations in the brain in ageing and neurodegeneration.

Astroglia or astrocytes, the most abundant cells in the brain, are interposed between neuronal synapses and the microvasculature in the brain’s gray matter. This unique anatomical location allows astroglia to play pivotal roles in brain metabolism as well as in the regulation of cerebral blood flow. In particular, astroglial cellular metabolic compartmentation exerts supportive roles in dedicating neurons to the generation of action potentials and protects neurons against the oxidative stress associated with their high energy consumption. Key products of astroglia include lactate and ketone bodies (beta-hydroxybutyrate and acetoacetate), which can also be produced avidly by muscle and liver, respectively. Therefore, brain cells, skeletal muscles, and hepatocytes constitute a metabolic compartmentation in the whole body. In this chapter, I will focus on brain cells, especially astroglia, since the impairment of normal astroglial function can lead to numerous neurological disorders including stroke, neurodegenerative diseases, and neuro-immunological diseases. I will also discuss the metabolic responses of brain cells in terms of food consumption and exercise. A better understanding of the astroglial metabolic response is expected to lead to the development of novel therapeutic strategies for diverse neurological diseases.

Endocrine abnormalities are very common in patients with chronic autoimmune rheumatic diseases (CARDs) due to the systemic involvement of the central nervous system and endocrine glands. In recent years, the response of the endocrine (and also neuronal) system to peripheral inflammation has been linked to overall energy regulation of the diseased body and bioenergetics of immune cells. In CARDs, hormonal and neuronal pathways are outstandingly important in partitioning energy-rich fuels from muscle, brain, and fat tissue to the activated immune system. Neuroendocrine regulation of fuel allocation has been positively selected as an adaptive programme for transient serious, albeit non-life-threatening, inflammatory episodes. In CARDs, mistakenly, the adaptive programmes are used again but for a much longer time leading to systemic disease sequelae with endocrine (and also neuronal) abnormalities. The major endocrine alterations are depicted in the following list: mild activation of the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system, inadequate secretion of ACTH and cortisol relative to inflammation, loss of androgens, inhibition of the hypothalamic-pituitary-gonadal axis and fertility problems, high serum levels of oestrogens relative to androgens, fat deposits adjacent to inflamed tissue, increase of serum prolactin, and hyperinsulinaemia (and the metabolic syndrome). Neuroendocrine abnormalities are demonstrated using this framework that can explain many CARD-related endocrine disturbances. This chapter gives an overview on pathophysiology of neuroendocrine alterations in the context of energy regulation.

The previous chapter dealt with the solution of the cable equation in response to current pulses and steps within a single unbranched cable. However, real nerve cells possess highly branched and extended dendritic trees with quite distinct morphologies. Figure 3.1 illustrates the fantastic variety of dendritic trees found throughout the animal kingdom, ranging from neurons in the locust to human brain cells and cells from many different parts of the nervous system. Some of these cells are spatially compact, such as retinal amacrine cells, which are barely one-fifth of a millimeter across, while some cells have immense dendritic trees, such as α motoneurones in the spinal cord extending across several millimeters. Yet, in all cases, neurons are very tightly packed: in vertebrates, peak density appears to be reached in the granule cell layer of the human cerebellum with around 5 million cells per cubic millimeter (Braitenberg and Atwood, 1958) while the packing density of the cells filling the 0.25 mm3 nervous system of the housefly Musca domestica is around 1.2 million cells per cubic millimeter (Strausfeld, 1976). The dendritic arbor of some cell types encompasses a spherical volume, such as for thalamic relay cells, while other cells, such as the cerebellar Purkinje cell, fill a thin slablike volume with a width less than one-tenth of their extent. Different cell types do not appear at random in the brain but are unique to specific parts of the brain. By far the majority of excitatory cells in the cortex are the pyramidal cells. Yet even within this class, considerable diversity exists. But why this diversity of shapes? To what extent do these quite distinct dendritic architectures reflect differences in their roles in information processing and computation? What influence does the dendritic morphology have on the electrical properties of the cell, or, in other words, what is the relationship between the morphological structure of a cell and its electrical function? One of the few cases where a quantitative relationship between form and some aspect of neuronal function has been established is the retinal neurons.

Current treatments of neurological and neurodegenerative diseases are limited due to the lack of a truly non-invasive, transient, and regionally selective brain drug delivery method. The brain is particularly difficult to deliver drugs to because of the blood-brain barrier (BBB). The impermeability of the BBB is due to the tight junctions connecting adjacent endothelial cells and highly regulatory transport systems of the endothelial cell membranes. The main function of the BBB is ion and volume regulation to ensure conditions necessary for proper synaptic and axonal signaling. However, the same permeability properties that keep the brain healthy also constitute the cause of the tremendous obstacles posed in its pharmacological treatment. The BBB prevents most neurologically active drugs from entering the brain and, as a result, has been isolated as the rate-limiting factor in brain drug delivery. Until a solution to the trans-BBB delivery problem is found, treatments of neurological diseases will remain impeded. Over the past decade, methods that combine Focused Ultrasound (FUS) and microbubbles have been shown to offer the unique capability of noninvasively, locally and transiently opening the BBB so as to treat central nervous system (CNS) diseases. Four of the main challenges that lie ahead are to: 1) assess its safety profile, 2) unveil the mechanism by which the BBB opens and closes, 3) control and predict the opened BBB properties and duration of the opening and 4) assess its premise in brain drug delivery. All these challenges will be discussed, findings in both small (mice) and large (non-human primates) animals will be shown and finally the case for this technique for clinical applications will be made.

In order to expand on potential injury mechanisms to the brain, a micromechanical structural representation of the gray matter must be developed. The gray matter contains a high volume of capillary vasculature that supplies the necessary oxygen required for maintaining healthy cell and brain function. Even short disruptions in this blood supply and the accompanying dissolved oxygen can lead to neuronal cell damage and death. It has been shown that increased shearing forces within the blood, such as those found near stents and artificial heart valves, can lead to platelet activation and aggregation, causing clots to form and potential disruptions in blood flow and oxygen distribution. Current macro-scale computational brain modeling can incorporate the larger main vasculature of the brain, but it becomes too computationally expensive to incorporate the smaller vessels. These larger scale models can be used to reveal how forces to the head are transmitted down to a scale slightly larger than the smallest capillaries within the gray matter. In order to investigate the response and potential damage to capillaries and platelets within the brain, a micromechanical computational model is developed incorporating the gray matter, capillaries, and blood, which is composed of plasma, red blood cells, and platelets. The red blood cells are a necessary component for the model for damage as it comprises almost half of the volume of blood and is the major contributor to the non-Newtonian behavior. The model combines both fluids and viscoelastic solid materials (the gray matter and the vascular wall). The deviatoric stress, strain and strain rate of the platelets in response to an externally applied load is measured and will determine the potential for platelet aggregation and clot formation. The micromechanical model is also used to provide verification and refinement for existing constitutive models for the gray matter used in meso- and macro-scale computational models.

Cerebral arteries play an important role in the regulation of cerebral blood flow through autoregulation, a well established phenomenon which is caused by a combination of myogenic, neuronal and metabolic mechanisms [1]. Myogenic reactivity is the ability of the vascular smooth muscle cells (SMC) to contract in response to stretch or to an increase in transmural pressure (TMP), and to dilate in response to a decrease in TMP [2]. It is this active constriction of arteries within the autoregulatory range that prompts studies of not just passive mechanical properties, but also active mechanical properties. Passive properties provide an understanding of the behavior of the extracellular matrix components of arteries (i.e. collagen and elastin); but, in order to understand how the artery behaves in vivo, it is necessary to understand the mechanical properties with smooth muscle cell activation. Mechanical properties might also be altered if the vessel is diseased or damaged. Ischemia has been shown to reduce vascular tone, which might lead to brain tissue damage during stroke [3]. Therefore studying the mechanical properties of vessels in disease states to determine if they are able to adequately take part in controlling local blood flow is also important.