Árpád Dobolyi

Astrocytes convert network excitation to tonic inhibition of neurons

BackgroundGlutamate and γ-aminobutyric acid (GABA) transporters play important roles in balancing excitatory and inhibitory signals in the brain. Increasing evidence suggest that they may act concertedly to regulate extracellular levels of the neurotransmitters.ResultsHere we present evidence that glutamate uptake-induced release of GABA from astrocytes has a direct impact on the excitability of pyramidal neurons in the hippocampus. We demonstrate that GABA, synthesized from the polyamine putrescine, is released from astrocytes by the reverse action of glial GABA transporter (GAT) subtypes GAT-2 or GAT-3. GABA release can be prevented by blocking glutamate uptake with the non-transportable inhibitor DHK, confirming that it is the glutamate transporter activity that triggers the reversal of GABA transporters, conceivably by elevating the intracellular Na+ concentration in astrocytes. The released GABA significantly contributes to the tonic inhibition of neurons in a network activity-dependent manner. Blockade of the Glu/GABA exchange mechanism increases the duration of seizure-like events in the low-[Mg2+] in vitro model of epilepsy. Under in vivo conditions the increased GABA release modulates the power of gamma range oscillation in the CA1 region, suggesting that the Glu/GABA exchange mechanism is also functioning in the intact hippocampus under physiological conditions.ConclusionsThe results suggest the existence of a novel molecular mechanism by which astrocytes transform glutamatergic excitation into GABAergic inhibition providing an adjustable, in situ negative feedback on the excitability of neurons.

The neuroprotective functions of transforming growth factor beta proteins.

Transforming growth factor beta (TGF-β) proteins are multifunctional cytokines whose neural functions are increasingly recognized. The machinery of TGF-β signaling, including the serine kinase type transmembrane receptors, is present in the central nervous system. However, the 3 mammalian TGF-β subtypes have distinct distributions in the brain suggesting different neural functions. Evidence of their involvement in the development and plasticity of the nervous system as well as their functions in peripheral organs suggested that they also exhibit neuroprotective functions. Indeed, TGF-β expression is induced following a variety of types of brain tissue injury. The neuroprotective function of TGF-βs is most established following brain ischemia. Damage in experimental animal models of global and focal ischemia was shown to be attenuated by TGF-βs. In addition, support for their neuroprotective actions following trauma, sclerosis multiplex, neurodegenerative diseases, infections, and brain tumors is also accumulating. The review will also describe the potential mechanisms of neuroprotection exerted by TGF-βs including anti-inflammatory, -apoptotic, -excitotoxic actions as well as the promotion of scar formation, angiogenesis, and neuroregeneration. The participation of these mechanisms in the neuroprotective effects of TGF-βs during different brain lesions will also be discussed.

Glutamate Uptake Triggers Transporter-Mediated GABA Release from Astrocytes

Background Glutamate (Glu) and γ-aminobutyric acid (GABA) transporters play important roles in regulating neuronal activity. Glu is removed from the extracellular space dominantly by glial transporters. In contrast, GABA is mainly taken up by neurons. However, the glial GABA transporter subtypes share their localization with the Glu transporters and their expression is confined to the same subpopulation of astrocytes, raising the possibility of cooperation between Glu and GABA transport processes. Methodology/Principal Findings Here we used diverse biological models both in vitro and in vivo to explore the interplay between these processes. We found that removal of Glu by astrocytic transporters triggers an elevation in the extracellular level of GABA. This coupling between excitatory and inhibitory signaling was found to be independent of Glu receptor-mediated depolarization, external presence of Ca2+ and glutamate decarboxylase activity. It was abolished in the presence of non-transportable blockers of glial Glu or GABA transporters, suggesting that the concerted action of these transporters underlies the process. Conclusions/Significance Our results suggest that activation of Glu transporters results in GABA release through reversal of glial GABA transporters. This transporter-mediated interplay represents a direct link between inhibitory and excitatory neurotransmission and may function as a negative feedback combating intense excitation in pathological conditions such as epilepsy or ischemia.

Calcitonin Gene-Related Peptide- Containing Pathways in the Rat Forebrain

The present study focuses on the topographical distribution of calcitonin gene‐related peptide (CGRP)‐containing cell bodies and fibers and their connections and pathways in the rat forebrain. We confirm previously reported CGRP projections from the perifornical area of the hypothalamus to the lateral septum, from the posterior thalamus to the caudate putamen and cerebral cortex, and from the parabrachial nuclei to the central extended amygdala, lateral hypothalamus, and ventromedial thalamus. Despite previous descriptions of CGRP in the central nervous system, important neuroanatomical aspects of the forebrain CGRP system remained obscure, which we addressed by using brain lesion techniques combined with modern immunohistology. We first report CGRP terminal fields in the olfactory‐anterior septal region and also CGRP projections from the parabrachial nuclei to the olfactory‐anterior septal region, the medial prefrontal cortex, the interstitial nucleus of the anterior commissure, the nucleus of the lateral olfactory tract, the anterior amygdaloid area, the posterolateral cortical amygdaloid nucleus, and the dorsolateral part of the lateral amygdaloid nucleus. In addition, we identified a CGRP cell group in the premamillary nuclei and showed that it projects to the medial CGRP layer of the lateral septum. CGRP fibers usually join other pathways rather than forming bundles. They run along the fornix from the hypothalamus, along the supraoptic decussations or the inferior thalamic peduncle‐stria terminalis pathway from the posterior thalamus, and along the superior cerebellar peduncle, thalamic fasciculus, and ansa peduncularis from the parabrachial nuclei. This description of the forebrain CGRP system will facilitate investigation of its role in higher brain functions. J. Comp. Neurol. 489:92–119, 2005.

Anatomical and physiological evidence for involvement of tuberoinfundibular peptide of 39 residues in nociception.

The parathyroid hormone 2 (PTH2) receptors anatomical distribution suggests that, among other functions, it may be involved in modulation of nociception. We localized PTH2 receptor protein to spinal cord lamina II and showed that it is synthesized by subpopulations of primary sensory neurons and intrinsic spinal cord dorsal horn neurons. Tuberoinfundibular peptide of 39 residues (TIP39) selectively activates the PTH2 receptor. Intraplantar microinjection of TIP39 caused a paw-withdrawal response and intrathecal injection caused scratching, biting, and licking, a nocifensive response. Intrathecal administration of a TIP39 antibody decreased sensitivity in tail-flick and paw-pressure assays. Intrathecal administration of TIP39 potentiated responses in these assays. We determined the sequence of TIP39s precursor and found that mRNA encoding TIP39 and TIP39-like immunoreactivity is concentrated in two brainstem areas, the subparafascicular area and the caudal paralemniscal nucleus. Cells in these areas project to the superficial dorsal horn of the spinal cord. Our data suggest that TIP39 released from supraspinal fibers potentiates aspects of nociception within the spinal cord.

Expression and distribution of tuberoinfundibular peptide of 39 residues in the rat central nervous system.

Tuberoinfundibular peptide of 39 residues (TIP39) has been recently purified and identified as a selective ligand for the parathyroid hormone 2 receptor. As a next step toward understanding its functions, we report the expression and distribution of TIP39 in the rat central nervous system. In situ hybridization histochemistry and immunocytochemistry revealed TIP39‐containing cell bodies in three distinct areas. The major one comprises the subparafascicular area posterior through the intralaminar nucleus of the thalamus; a second is the medial paralemniscal nucleus at the pontomesencephalic junction; and a third is in the dorsal and dorsolateral hypothalamic areas, which contained a few, scattered cell bodies. We found, in contrast to the highly restricted localization of TIP39‐containing cell bodies, a much more widespread localization of TIP39‐containing fibers. The highest density of fibers was observed in limbic areas such as the septum, the amygdala, and the bed nucleus of the stria terminalis; in areas involved in endocrine regulation, such as the hypothalamic dorsomedial, paraventricular, periventricular, and arcuate nuclei; in auditory areas, such as the ectorhinal and temporal cortices, inferior colliculus, medial geniculate body, and some of the nuclei of the superior olivary complex; and in the dorsolateral funiculus of the spinal cord. The localization of TIP39‐containing nuclei and fibers provides an anatomical basis for previously demonstrated endocrine and nociceptive effects of TIP39 and suggests additional functions for TIP39, one apparent candidate being the regulation of auditory information processing. J. Comp. Neurol. 455:547–566, 2003. Published 2002 Wiley‐Liss, Inc.

Analysis of purine and pyrimidine bases, nucleosides and deoxynucleosides in brain microsamples (microdialysates and micropunches) and cerebrospinal fluid

A new chromatographic method is reported for the synchronous analysis of endogenous purine and pyrimidine bases, ribonucleosides, and deoxyribonucleosides in brain samples. An optimized gradient chromatography system with a cooled reversed-phase column allows the detection of these compounds in very low concentrations in microsamples (microdialysates and micropunches). Chromatographic peaks were identified via the retention times of known standards, with detection at two wavelengths, and also by electrospray tandem mass spectrometry, which permits the identification of certain compounds at extremely low concentrations. The method was tested on in vivo brain microdialysis samples, micropunch tissue sample and cerebrospinal fluid of rats. Extracellular concentrations of pyrimidine metabolites in brain samples and of various purine metabolites in thalamic samples are reported here first. A comparison of the results on microdialysis and cerebrospinal fluid samples suggests that the analysis of cerebrospinal fluid provides limited information on the local extracellular concentrations of these compounds. Basic dialysis experiments revealed temporarily stable baseline levels one hour after implantation of the microdialysis probes. An elevated potassium concentration in the perfusion solution caused increases in the extracellular levels of adenosine and its metabolites, and of guanosine and the pyrimidine nucleoside uridine.

Neurons containing tuberoinfundibular peptide of 39 residues project to limbic, endocrine, auditory and spinal areas in rat.

Accumulating evidence suggests that tuberoinfundibular peptide of 39 residues (TIP39) may be the endogenous ligand of the parathyroid hormone 2 receptor. The vast majority of TIP39-containing neurons are localized in two regions, the subparafascicular area at the thalamic-midbrain junction, and the medial paralemniscal nucleus in the rostral pons. In contrast to the restricted localization of TIP39-containing cell bodies, TIP39-containing fibers have a widespread distribution. TIP39 neurons were lesioned electrolytically to determine the origin of TIP39-containing fibers within different parts of the rat CNS. Following bilateral lesions of the medial subparafascicular area including the subparafascicular nucleus, TIP39-immunoreactive fibers almost completely disappeared from forebrain regions including the anterior limbic cortical areas, the shell and cone portions of the nucleus accumbens, the lateral septum, the bed nucleus of the stria terminalis, the amygdaloid nuclei, the fundus striati, the subiculum, the thalamic paraventricular nucleus, and the hypothalamic paraventricular, dorsomedial and arcuate nuclei. Unilateral lesions of the medial and the lateral subparafascicular area demonstrated that the projections are ipsilateral and that medial lesions produce higher reductions in the density of TIP39 fibers except in the amygdala and the hypothalamus. Following lesions of the medial paralemniscal nucleus, TIP39-immunoreactive fibers disappeared from the medial geniculate body, the periaqueductal gray, the deep layers of the superior colliculus, the external cortex of the inferior colliculus, the cuneiform nucleus, the nuclei of the lateral lemniscus, the lateral parabrachial nucleus, the locus coeruleus, the subcoeruleus area, the medial nucleus of the trapezoid body, the periolivary nuclei, and the spinal cord, suggesting that these regions receive TIP39-containing fibers from the medial paralemniscal nucleus, and unilateral lesions demonstrated that the projections are ipsilateral. The projections of the TIP39-containing cells in the subparafascicular area suggest their involvement in limbic and endocrine functions, while the projections of the TIP39-containing cells in the medial paralemniscal nucleus suggest their involvement in auditory and nociceptive functions.

Distribution of mRNAs encoding transforming growth factors-β1, -2, and -3 in the intact rat brain and after experimentally induced focal ischemia

Transforming growth factors‐β1 (TGF‐β1), ‐2, and ‐3 form a small group of related proteins involved in the regulation of proliferation, differentiation, and survival of various cell types. Recently, TGF‐βs were also demonstrated to be neuroprotective. In the present study, we investigated their distribution in the rat brain as well as their expression following middle cerebral artery occlusion. Probes were produced for all types of TGF‐βs, and in situ hybridization was performed. We demonstrated high TGF‐β1 expression in cerebral cortex, hippocampus, central amygdaloid nucleus, medial preoptic area, hypothalamic paraventricular nucleus, substantia nigra, brainstem reticular formation and motoneurons, and area postrema. In contrast, TGF‐β2 was abundantly expressed in deep cortical layers, dentate gyrus, midline thalamic nuclei, posterior hypothalamic area and mamillary body, superior olive, areas of monoaminergic neurons, spinal trigeminal nucleus, dorsal vagal complex, cerebellum, and choroid plexus, and a high level of TGF‐β3 mRNA was found in cerebral cortex, hippocampus, basal amygdaloid nuclei, lateral septal nucleus, several thalamic nuclei, arcuate and supramamillary nuclei, superior colliculus, superior olive, brainstem reticular formation and motoneurons, area postrema, and inferior olive. Focal brain ischemia induced TGF‐βs with markedly different expression patterns. TGF‐β1 was induced in the penumbral region of cortex and striatum, whereas TGF‐β2 and ‐β3 were induced in different layers of the ipsilateral cortex. The expression of the subtypes of TGF‐βs in different brain regions suggests that they are involved in the regulation of different neurons and bind to different latent TGF‐β binding proteins. Furthermore, they might have subtype‐specific functions following ischemic attack. J. Comp. Neurol. 518:3752–3770, 2010.

Tuberoinfundibular Peptide of 39 Residues Is Activated during Lactation and Participates in the Suckling-Induced Prolactin Release in Rat

Tuberoinfundibular peptide of 39 residues (TIP39) and the PTH-2 receptor (PTH2R) constitute a peptide-receptor neuromodulator system. Based on the abundance of TIP39 fibers and axonal terminals as well as PTH2R-containing neurons and their processes in the hypothalamic para- and periventricular and arcuate nuclei TIP39 has been suggested to play a role in neuroendocrine regulation. We showed previously that TIP39 expression decreased dramatically by adulthood. In the present study, using in situ hybridization histochemistry, real-time RT-PCR, and immunohistochemistry, we found that TIP39 mRNA and peptide expression levels are markedly elevated in the posterior intralaminar complex of the thalamus (PIL) of lactating dams, one of the three locations of TIP39-containing cell bodies in the brain. In addition, in mother rats, these TIP39 neurons showed Fos expression in response to pup exposure. Transection of TIP39 fibers originating in the PIL resulted in an ipsilateral disappearance of TIP39 immunoreactivity throughout the mediobasal hypothalamus of mother rats, suggesting that TIP39 fibers there arise from the PIL. To elucidate the function of TIP39 activation in dams, mothers separated from their pups for 4 h on postpartum d 9 received injection of a PTH2R antagonist into the lateral ventricle 5 min before returning the pups. Blood samples were taken seven times during the experimental period through jugular cannulae. The PTH2R antagonist administered in two different concentrations markedly inhibited suckling-induced elevation of plasma prolactin levels in a dose-dependent manner. These results suggest that TIP39 neurons in the PIL may regulate suckling-induced prolactin release in rat dams.