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H Kettenmann

Publications and source records attributed to H Kettenmann.

At least 37 records · Page 2Linked to original sources
PubMed2000

Distinct physiologic properties of microglia and blood-borne cells in rat brain slices after permanent middle cerebral artery occlusion.

The authors investigated the time course of leukocyte infiltration compared with microglial activation in adult rat brain slices after permanent middle cerebral artery occlusion (MCAO). To distinguish peripheral leukocytes from microglia, the blood cells were prelabeled in vivo with Rhodamine 6G (Rhod6G) i.v. before induction of ischemia. At specific times after infarct, invading leukocytes, microglia, and endothelial cells were labeled in situ with isolectin (IL)B4-FITC (ILB4). Six hours after MCAO only a few of the ILB4+ cells were colabeled by Rhod6G. These cells expressed the voltage-gated inwardly and outwardly rectifying K+ currents characteristic of macrophages. The majority of the ILB4+ cells were Rhod6G- and expressed a lack of voltage-gated channels, recently described for ramified microglial cells in brain slices, or exhibited only an inward rectifier current, a unique marker for cultured (but unstimulated) microglia. Forty-eight hours after MCAO, all blood-borne and the majority of Rhod6G- cells expressed outward and inward currents indicating that the intrinsic microglial population exhibited physiologic features of stimulated, cultured microglia. The ILB4+/Rhod6G- intrinsic microglial population was more abundant in the border zone of the infarct and their morphology changed from radial to ameboid. Within this zone, the authors observed rapidly migrating cells and recorded this movement by time-lapse microscopy. The current findings indicate that microglial cells acquire physiologic features of leukocytes at a later time point after MCAO.

Animals↗
PubMed1999

Heterogeneous expression of voltage-gated potassium channels of the shaker family (Kv1) in oligodendrocyte progenitors.

Outwardly rectifying K(+) channels determine the membrane conductance and influence the proliferation rate of glial progenitor cells. To analyze the molecular identity and the functional role of K(+) channels in glial progenitors of mouse brain, expression of shaker-type Kv1 genes was studied at three levels: (1) presence of Kv1 mRNAs, (2) biosynthesis of channel proteins and (3) electrophysiological and pharmacological properties of K(+) currents. mRNA expression of Kv1.1 to Kv1.6 genes was studied by single-cell reverse transcription-mediated polymerase chain reaction (RT-PCR) using degenerate primers to amplify the six Kv1 transcripts. Most cells expressed several mRNA combinations simultaneously. In more than half of the cells, messages for Kv1.2, Kv1.5 and Kv1.6 were found, while Kv1.1, Kv1.3 and Kv1.4 were detected in only a minority of cells. In contrast, at the level of protein expression - employing immunocytochemistry with subtype-specific antibodies - Kv1. 2 and Kv1.3 were undetectable (<2%), while almost all cells expressed Kv1.4 (85%), Kv1.5 (99%) and Kv1.6 (99%). Kv1.1 was present in a minor cell population (10%). Functional contribution of Kv1 proteins to progenitor membrane conductance was determined by analyzing the voltage-dependence of K(+) current activation and inactivation as well as their current sensitivities to the subtype-preferring blockers and toxins tetraethylammonium (TEA), 4-aminopyridine (4-AP), charybdotoxin (CTX), alpha-dendrotoxin (DTX) and mast-cell degranulating peptide (MCDP). From these results, it is concluded: first, glial progenitor cells can express all transcripts of the six Kv1 genes, but do not express all proteins; second, Kv1.4, Kv1.5 and Kv1.6 proteins are most abundant and were found in the majority of cells; and third, K(+) currents flow predominantly either through heteromeric channel complexes or through homomeric Kv1.5 ion pores, but not through homomeric Kv1.4 or Kv1.6 channels.

Animals↗
PubMed1999

Induction of potassium channels in mouse brain microglia: cells acquire responsiveness to pneumococcal cell wall components during late development.

Lipopolysaccharides derived from cell walls of Gram-negative bacteria have proven a useful tool to simulate bacterial infection of the central nervous system. Rapid activation of microglia within the brain parenchyma as well as in vitro has thereby been shown to be an early event upon bacterial or lipopolysaccharide challenges. Less is known about microglial responses to a contact with Gram-positive bacteria, such as Streptococcus pneumoniae, a lethal pathogen causing meningitis with a 30% mortality rate. In the present study, we compared lipopolysaccharide-induced microglial activation in vitro with that induced by preparations of pneumococcal cell walls. As a readout of microglial activation, we studied by patch-clamp recording the expression of outward rectifying potassium currents (IK+OR), which are known to be induced by lipopolysaccharide. We found that pneumococcal cell walls and lipopolysaccharide induced a similar type of IK+OR. Stimulation of IK+OR by pneumococcal cell walls and lipopolysaccharide involved protein synthesis since it was not induced in the presence of cycloheximide. Pharmacological characterization of the pneumococcal cell wall- and lipopolysaccharide-induced currents with specific ion channel blockers indicated for both cases expression of the charybdotoxin/margatoxin-sensitive Kv1.3 subtype of the Shaker family of voltage-dependent potassium channels. Activation of the outward currents by pneumococcal cell walls depended on the developmental stage: while lipopolysaccharide triggered IK+OR in both embryonal and postnatal microglial cells, pneumococcal cell walls had only a marginal effect on embryonal cells. This, however, does not imply that embryonic microglial cells are unresponsive to pneumococcal cell walls. In both embryonic and postnatal cells, (i) the amplitude of the constitutively expressed inward rectifying potassium current was significantly reduced, (ii) tumor necrosis factor-a was released and (iii) the cells changed their morphology, similarly as it was induced by lipopolysaccharide treatment. Thus, embryonic microglial cells are sensitive to pneumococcal cell wall challenges, but respond with a distinctly different pattern of physiological reactions. The expression of IK+OR could thus be a suitable tool to study signalling cascades selectively involved in the activation of microglia by Gram-negative and -positive cell wall components and to functionally distinguish between populations of microglial cells.

Animals↗
PubMed1999

Glutamate-triggered calcium signalling in mouse bergmann glial cells in situ: role of inositol-1,4,5-trisphosphate-mediated intracellular calcium release.

The mechanisms of glutamate-induced changes in intracellular free calcium concentration in Bergmann glial cells in mouse cerebellar slices were investigated by Fura-2-based microfluorimetry. Extracellular applications of glutamate, quisqualate and kainate triggered an increase in cytoplasmic calcium concentration, whereas N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate were ineffective. The calcium elevation triggered by kainate was completely blocked by removal of calcium ions from the external solutions or by slice incubation with 6-cyano-7-nitroquinoxaline-2,3-dione. Conversely, both glutamate- and quisqualate-induced intracellular calcium transients were only slightly attenuated by slice incubation with either 6-cyano-7-nitroquinoxaline-2,3-dione or calcium-free solution, suggesting the intracellular origin for calcium ions. The glutamate-triggered cytosolic calcium increases were inhibited by slice incubation with thapsigargin, the inhibitor of intracellular calcium pumps, or by intracellular perfusion of Bergmann glial cells with heparin, the antagonist of inositol-1,4,5-trisphosphate-gated calcium release channels. Therefore the calcium release from inositol-1,4,5-trisphosphate-sensitive intracellular stores plays the major role in glutamate-induced calcium signalling. We concluded that Bergmann glial cells express calcium permeable ionotropic glutamate receptors, which might be important for generation of fast calcium signals. However, slow glutamate-evoked calcium signals are mostly determined by inositol-1,4,5-trisphosphate-dependent intracellular signalling chain.

Amino Acids↗
PubMed1999

Microdomains for neuron-glia interaction: parallel fiber signaling to Bergmann glial cells.

Astrocytes are considered a reticulate network of cells, through which calcium signals can spread easily. In Bergmann glia, astrocytic cells of the cerebellum, we identified subcellular compartments termed 'glial microdomains'. These elements have a complex surface consisting of thin membrane sheets, contain few mitochondria and wrap around synapses. To test for neuronal interaction with these structures, we electrically stimulated parallel fibers. This stimulation increased intracellular calcium concentration ([Ca2+]i) in small compartments within Bergmann glial cell processes similar in size to glial microdomains. Thus, a Bergmann glial cell may consist of hundreds of independent compartments capable of autonomous interactions with the particular group of synapses that they ensheath.

Animals↗
PubMed1999

Microglial activation by components of gram-positive and -negative bacteria: distinct and common routes to the induction of ion channels and cytokines.

Gram-positive Streptococcus pneumoniae is the major pathogen causing lethal meningitis in adults. We used pneumococcal cell walls (PCW) to investigate microglial consequences of a bacterial challenge and to determine the role of serum in the activation process. PCW caused the characteristic induction of an outwardly rectifying K+ channel (IK+(OR)), together with a concomitant suppression of the constitutively expressed inward rectifier K+ current, and evoked the release of tumor necrosis factor-alpha (TNF alpha), interleukin-6 (IL-6), IL-12, KC, macrophage inflammatory protein (MIP) 1alpha and MIP-2. Serum presence strongly facilitated the PCW effects, similarly as observed for lipopolysaccharide (LPS) from gram-negative Escherichia coli. The inflammatory cytokine, interferon-gamma (IFNgamma) induced the same electrophysiological changes, but independent of serum. Recombinant LPS binding protein (LBP) could partially replace serum activity in LPS stimulations. In contrast, neither LBP nor an antibody-mediated blockade of the LPS receptor, CD14 had significant influences on PCW-inducible changes. Cell surface interactions and cofactor involvement in microglial activation by gram-positive bacteria are thus distinct from the mechanisms employed by LPS. Moreover, tyrphostin AG126, a protein kinase inhibitor that prevents activation of the mitogen-activated protein kinase, p42MAPK (ERK2), potently blocked the PCW-stimulated cytokine release while having only a limited effect on LPS-inducible cytokines. In contrast, AG126 did not influence IK+(OR) inductions. This indicates that PCW recruits more than 1 intracellular signaling pathway to trigger the various responses and that different bacterial agents signal through both common and individual routes during microglial activation.

Acute-Phase Proteins↗
PubMed1998

Alternative splicing of mouse IL-15 is due to the use of an internal splice site in exon 5.

IL-15 is a pleiotropic cytokine modulating growth and differentiation of several hematopoietic cell types. Recently, we have demonstrated that mouse microglial cells, the brain macrophages, express both IL-15 and IL-15/IL-2 receptors. Based on single-cell RT-PCR data, we describe here an alternatively spliced IL-15 mRNA variant found in a small subpopulation of mouse microglia (5%, 3 out of 60 cells expressing IL-15 transcripts). PCR cycle sequencing of this larger transcript revealed the mouse homologue of the alternatively spliced exon A as it is known from the human IL-15 gene. Analysis of the corresponding mouse IL-15 gene region shows that the larger IL-15 transcript contains an yet unidentified 5' sequence of exon 5 while the shorter transcript uses an internal splice acceptor site. The mouse exon 5A segment has a length of 136 nt (17 nt longer than the human exon A). It contains five in-frame stop codons at its 5' end and a new translation initiation site at its 3' end. This new start site is surrounded by a favourable Kozak consensus sequence suggesting a more efficient translation rate. Further translational control by stem-loop binding factors is inferred by a predicted RNA stem-loop structure around the start site. Insertion of exon 5A would lead to an IL-15 polypeptide with a shortened leader sequence of 26 amino acids, as compared to the 48 amino acid leader sequence encoded by the transcript lacking exon 5A. Thus, the final IL-15 protein of the two splice variants is identical; different leader sequences could, however, lead to differences in the intracellular sorting, processing and/or secretion of IL-15.

Alternative Splicing↗
PubMed1998

Long-term activation of capacitative Ca2+ entry in mouse microglial cells.

The cytoplasmic free calcium concentration ([Ca2+]i) was measured in cultured microglial cells with the Ca2+-sensitive fluorescent dye Fura-2 using a digital imaging system. Stimulation of P2 purinergic receptors by ATP or UTP always evoked a [Ca2+]i elevation. The ATP-induced Ca2+ response involved both Ca2+ influx through ionotropic receptors and Ca2+ release from intracellular pools, whereas UTP selectively stimulated intracellular Ca2+ release. When intracellular Ca2+ release was stimulated in the absence of extracellular Ca2+, the readmission of extracellular Ca2+ caused a large rebound [Ca2+]i increase. Following this rebound, [Ca2+]i did not return to the initial resting level, but remained for long periods of time (up to 20 min), at a new, higher steady-state level. Both the amplitude of the rebound Ca2+ transient and the new plateau level strongly correlated with the degree of intracellular Ca2+ depletion, indicating the activation of a store-operated Ca2+ entry pathway. The elevated steady-state [Ca2+]i level was associated with a significant increase in the plasma membrane permeability to Ca2+, as changes in extracellular Ca2+ were reflected in almost immediate changes of [Ca2+]i. Similarly, blocking plasma-lemmal Ca2+ channels with the non-specific agonist La3+ (50 microM) caused a decrease in [Ca2+]i, despite the continuous presence of Ca2+ ions in the extracellular medium. After the establishment of the new, elevated steady-state [Ca2+]i level, stimulation of P2U metabotropic purinoreceptors did not induce a [Ca2+]i response. In addition, application of either thapsigargin (1 microM) or carbonyl cyanide chlorophenyl hydrazone (10 microM) failed to affect [Ca2+]i. We conclude that the maximal depletion of intracellular Ca2+ stores in mouse brain microglia determines the long-term activation of a plasma membrane Ca2+ entry pathway. This activation appears to be associated with a significant decrease in the capability of the intracellular Ca2+ stores to take up cytosolic Ca2+ once they have been maximally depleted.

Adenosine Triphosphate↗
PubMed1998

Functional GABA(A) receptors on human glioma cells.

Glioma cells in acute slices and in primary culture, and glioma-derived human cell lines were screened for the presence of functional GABA(A) receptors. Currents were measured in whole-cell voltage clamp in response to gamma-aminobutyric acid (GABA). While cells from the most malignant glioma, the glioblastoma multiforme, did not respond to GABA, an inward current (under our experimental conditions with high Cl- concentration in the pipette) was induced in gliomas of lower grades, namely in 71% of oligodendroglioma cells and in 62% of the astrocytoma cells. Glioma cell lines did not express functional GABA(A) receptors, irrespective of the malignancy of the tumour they originate from. The currents elicited by application of GABA were due to activation of GABA(A) receptors; the specific agonist muscimol mimicked the response, the antagonists bicuculline and picrotoxin blocked the GABA-activated current and the benzodiazepine receptor agonist flunitrazepam augmented the GABA-induced current and the benzodiazepine inverse agonist DMCM decreased the GABA current. Cells were heterogeneous with respect to the direction of the current flow as tested in gramicidin perforated patches: in some cells GABA hyperpolarized the membrane, while in the majority it triggered a depolarization. Moreover, GABA triggered an increase in [Ca2+]i in the majority of the tumour cells due to the activation of Ca2+ channels. Our results suggest a link between the expression of GABA receptors and the growth of glioma cells as the disappearance of functional GABA(A) receptors parallels unlimited growth typical for malignant tumours and immortal cell lines.

Anti-Bacterial Agents↗
PubMed1998

Glutamate receptor activation can trigger electrical activity in human glioma cells.

Cells from major types of gliomas, i.e. oligodendrogliomas and glioblastomas, are able to generate action potentials upon a current injection similar to neurons (Patt et al. (1996) Neuroscience, 71, 601-611; Labrakakis et al. (1997b) J. Neuropath. Exp. Neurol., 56, 243-254. Here, we report that activation of ionotropic glutamate receptors by the selective agonist, kainate, or by glutamate itself, depolarized the tumour cells in culture and living slices from tumour tissue, and can elicit volleys of action potentials, as recorded with the patch-clamp technique. Sixty-six percent of the glioblastoma cells, 44% of the astocytoma and 86% of the oligodendroglioma cells responded to glutamate and the specific agonist of AMPA/kainate receptors, kainate. The involvement of non-NMDA (N-methyl-D-aspartate) receptors is further supported by the observation that both kainate and glutamate currents were blocked by CNQX (6-cyano-7-nitroquinoxaline-2,3-dione). The receptor activation was accompanied by an increase in cytosolic Ca2+, as recorded with a fura-2 microfluorometric system. The Ca2+ elevation was mediated by the activation of Ca2+ channels due to membrane depolarization. The presence of voltage-gated Ca2+ channels was confirmed by patch-clamp experiments. Taken together, these findings imply that the electrophysiological properties of glioma cells are more reminiscent of those of neurons than of glial cells.

Calcium↗
PubMed1998

Oligodendrocytes and microglia are selectively vulnerable to combined hypoxia and hypoglycemia injury in vitro.

The major classes of glial cells, namely astrocytes, oligodendrocytes, and microglial cells were compared in parallel for their susceptibility to damage after combined hypoxia and hypoglycemia or hypoxia alone. The three glial cell types were isolated from neonatal rat brains, separated, and incubated in N2/CO2-gassed buffer-containing glucose or glucose substitutes, 2-deoxyglucose or mannitol (both nonmetabolizable sugars). The damage to the cells after 6 hours' exposure was determined at 0, 1, 3, 7 days based on release of lactate dehydrogenase and counting of ethidium bromide-stained dead cells, double-stained with cell-type specific markers. When 2-deoxyglucose replaced glucose during 6 hours of hypoxia, both oligodendrocytes and microglia rarely survived (18% and 12%, respectively). Astroglia initially increased the release of lactate dehydrogenase but maintained 98% to 99% viability. When mannitol, a radical scavenger and osmolarity stabilizer, replaced glucose during 6 hours of hypoxia, oligodendrocytes rarely survived (10%), astroglia survival remained at 99%, but microglia survival increased to 50%. After exposure to 6 and 42 hours, respectively, of hypoxic conditions alone, oligodendrocytes exhibited 10% survival whereas microglia and astroglia were only temporarily stressed and subsequently survived. In conclusion, oligodendrocytes, then microglia, are the most vulnerable glial cell types in response to hypoxia or hypoglycemia conditions, whereas astrocytes from the same preparations recover.

Animals↗
PubMed1998

Single-cell characterization of endothelin system gene expression in the cerebellum in situ.

To evaluate the expression of components of the endothelin (ET) system in single Purkinje neurons and Bergmann glial cells in situ, patch-clamp recording was combined with a multiplex RT-PCR approach. Cerebellar slices were rapidly isolated from 20- to 28-day-old mice. Cells were characterized morphologically and electrophysiologically and cell contents were aspirated and immediately reverse-transcribed. The cDNA was used as a template in a multiplex PCR reaction containing primers specific for ET-1, ET-2, and ET-3, ET-converting enzyme 1 (ECE-1) and ECE-2, and ETA and ETB receptors. The resulting PCR products were used as templates in a second PCR reaction containing only one pair of nested primers. Specific single bands were obtained from positive cells, which was confirmed by DNA sequencing of the PCR products. Of the 25 Purkinje neurons assayed, 84% were positive for ECE-1 mRNA and 68% for ECE-2 mRNA. No ET and ETA receptor mRNAs were detected, and only one cell was positive for ETB receptor mRNA. In Bergmann glial cells, ETB receptor mRNA was predominant. A total of 68% of the 25 cells assayed were positive. Sixteen percent were positive for ETA receptor mRNA, 8% for ECE-1 mRNA, and 12% for ECE-2 mRNA. Again, no ET mRNAs were detected. These results confirm the role of the ETB receptor in Bergmann glial cells and provide evidence for expression of ECE-1 and ECE-2 in Purkinje neurons.

Animals↗
PubMed1998

Glial calcium: homeostasis and signaling function.

Glial cells respond to various electrical, mechanical, and chemical stimuli, including neurotransmitters, neuromodulators, and hormones, with an increase in intracellular Ca2+ concentration ([Ca2+]i). The increases exhibit a variety of temporal and spatial patterns. These [Ca2+]i responses result from the coordinated activity of a number of molecular cascades responsible for Ca2+ movement into or out of the cytoplasm either by way of the extracellular space or intracellular stores. Transplasmalemmal Ca2+ movements may be controlled by several types of voltage- and ligand-gated Ca(2+)-permeable channels as well as Ca2+ pumps and a Na+/Ca2+ exchanger. In addition, glial cells express various metabotropic receptors coupled to intracellular Ca2+ stores through the intracellular messenger inositol 1,4,5-triphosphate. The interplay of different molecular cascades enables the development of agonist-specific patterns of Ca2+ responses. Such agonist specificity may provide a means for intracellular and intercellular information coding. Calcium signals can traverse gap junctions between glial cells without decrement. These waves can serve as a substrate for integration of glial activity. By controlling gap junction conductance, Ca2+ waves may define the limits of functional glial networks. Neuronal activity can trigger [Ca2+]i signals in apposed glial cells, and moreover, there is some evidence that glial [Ca2+]i waves can affect neurons. Glial Ca2+ signaling can be regarded as a form of glial excitability.

Animals↗
PubMed1998

Dye coupling between spinal cord oligodendrocytes: differences in coupling efficiency between gray and white matter.

Oligodendrocytes express two gap junction proteins, connexin32 (Cx32) and Cx45. To test for functional coupling between oligodendrocytes, cells were filled with the (Cx32-permeable) dyes Lucifer Yellow (LY) and Neurobiotin. Cells in slices from rat spinal cord were dialyzed via the patch pipette containing the dye while recording with the patch-clamp technique. The dye-labeled cells were identified as oligodendrocytes by their characteristic pattern of membrane currents and by morphology. In gray matter, 18% of the injected cells (N = 94) were coupled to more than three adjacent cells (slices from postnatal day 1 to 19). In contrast, in white matter, the dye was restricted to the injected cell (N = 63 for Lucifer Yellow injection only; N = 11 for LY and Neurobiotin) indicating a lack of functional coupling. Immunolabeling of Cx32 in mature oligodendrocytes of white matter revealed that the gap junction protein is localized on the cell bodies and abaxonal processes which occupy non-overlapping territories. In immature white matter and gray matter, Cx32 is mostly concentrated in the somatic region of the cells. In addition to Cx32, we have obtained immunocytochemical data that oligodendrocytes can express Cx45 with a labeling pattern different from the Cx32 expression. Two alternative interpretations of the coupling data are discussed: 1) that the presence of Cx32 in mature white matter oligodendrocytes does not serve for communication between cells, but rather for communication within oligodendrocytes in the sense of autocellular coupling, or 2) that the glial syncytium is furnished with a high degree of functional rectification at the oligodendrocytic side.

Animals↗
PubMed1997

Microglial phagocytosis is modulated by pro- and anti-inflammatory cytokines.

Activation of microglial cells in neurological diseases involves proliferation and the induction of phagocytic and cytotoxic properties. We studied the effects of four different cytokines on microglial phagocytosis of latex beads to gain further insights into the signals modulating different aspects of microglial activity. Granulocyte/macrophage colony stimulating factor and tumor necrosis factor-alpha enhanced microglial phagocytic activity as measured by flow cytometry. A phagocytosis inhibiting effect was observed after preincubation with transforming growth factor-beta1 and interleukin-4. In conclusion, the activating and deactivating cytokines differentially regulate microglial phagocytic activity in vitro and might also play an important role in vivo in modulating microglial activation to keep the balance between the protective, defensive and destructive, chronic inflammatory properties of microglia.

Animals↗
PubMed1997

Mouse brain microglia express interleukin-15 and its multimeric receptor complex functionally coupled to Janus kinase activity.

The cytokine, interleukin (IL)-15, and the T cell growth factor, IL-2, exhibit a similar spectrum of immune effects and share the IL-2 receptor (IL-2R) subunits IL-2Rbeta and IL-2Rgamma for signaling in hematopoietic cells. Numerous neuroregulatory activities of IL-2 have been suggested, but its expression in the normal central nervous system (CNS) is apparently very low and regionally restricted. We show by RNA and protein detection that IL-15, its specific receptor molecule, IL-15Ralpha, and the signal-transducing receptor subunits, IL-2Rbeta and IL-2Rgamma, are constitutively present in various regions of the developing and adult mouse brain. We further demonstrate, also at the single-cell level, that IL-15 and the components for IL-15Ralpha/IL-2Rbetagamma receptors are expressed by microglia. Tyrosine phosphorylation data are presented showing that IL-15 signaling in microglia involves Janus kinase 1 activity. At doses of 0.1-10 ng/ml, IL-15 affected functional properties of these cells, such as the production of nitric oxide, and supported their growth in culture, suggestive of a role as an autocrine growth factor. Microglial IL-15 could thus play a pivotal role in the CNS and may participate in certain CNS and neuroendocrine functions previously ascribed to IL-2.

Animals↗
PubMed1997

Endothelin-induced calcium signaling in cultured mouse microglial cells is mediated through ETB receptors.

Microglial cells are the intrinsic immunocompetent cells of the central nervous system, which are activated by brain tissue damage. In this paper we investigated the ability of endothelins (ETs), which are potent vasoconstrictors, to induce intracellular calcium signals in cultured microglia cells. Both endothelin-1 and endothelin-3 increased intracellular Ca2+ concentration ([Ca2+]i). These [Ca2+]i transients were mimicked by BQ3020, an ETB receptor agonist and blocked by BQ788, a selective ETB antagonist, respectively. The calcium signals induced by the endothelins persisted in Ca(2+)-free media. Transcripts encoding the ETB receptor were detected in purified microglial cultures and cDNA fragments derived from ETB receptor mRNA were amplified from 9% of electrophysiologically characterized microglial cells by the use of single-cell RT-PCR.

Animals↗