Dietrich van Calker

Adenosine A2b receptors mediate an increase in interleukin (IL)-6 mRNA and IL-6 protein synthesis in human astroglioma cells.

Abstract: The cytokine interleukin (IL)‐6 has recently been demonstrated to play a role in the pathology of Alzheimers disease (AD). The mechanisms leading to increased IL‐6 levels in brains of AD patients are still unknown. Because in experimental animals ischemia increases both the level of cytokines and the extracellular concentrations of adenosine in the brain, we hypothesized that these two phenomena may be functionally connected and that adenosine might increase IL‐6 gene expression in the brain. Here we show that the mixed A1 and A2 agonist 5′‐(N‐ethylcarboxamido)adenosine (NECA) induces an increase in IL‐6 mRNA levels and protein synthesis in the human astrocytoma cell line U373 MG. The A1‐specific agonists R‐phenylisopropyladenosine and cyclopentyladenosine are much less potent, and the A2a‐specific agonist CGS‐21680 shows only marginal effects. Increased levels of mRNA are already found within 30 min after NECA treatment. The A2a‐selective antagonists 8‐(3‐chlorostyryl)caffeine and KF17837 [(E)‐8‐(3,4‐dimethoxystyryl)‐1,3‐dipropyl‐7‐methylxanthine], which have also some antagonistic properties at A2b receptors, and the nonspecific adenosine antagonist 8‐phenyltheophylline were equipotent at inhibiting the NECA‐induced increase in IL‐6 protein synthesis, whereas the specific A1 antagonist 8‐cyclopentyl‐1,3‐dipropylxanthine is much less potent. The results indicate that adenosine A2b receptors participate in the regulation of the IL‐6 gene in astrocytoma cells.

CARBAMAZEPINE DISTINGUISHES BETWEEN ADENOSINE RECEPTORS THAT MEDIATE DIFFERENT SECOND MESSENGER RESPONSES

The mechanism of the therapeutic and prophylactic effects of carbamazepine (CBZ) in affective psychoses is unknown but may in part be related to the potent competitive interaction of CBZ with adenosine-binding sites in the brain. The anticonvulsant and sedative properties of CBZ are reminiscent of the effects evoked by adenosine-agonists and contrast sharply with the opposite actions of adenosine-antagonists like caffeine. However, indirect evidence suggests an antagonist- rather than an agonist-like activity of CBZ at adenosine-receptors. We have used various model systems, in which adenosine receptor subtypes mediate different second messenger-responses, to investigate this-apparent paradox. CBZ was found to antagonize the A1-receptor-mediated inhibition of cyclic AMP accumulation in cultured astroblasts and in GH3-cells. Furthermore, CBZ also inhibits the adenosine-induced increase in the level of cyclic AMP in cultured astroblasts, which is mediated by low-affinity A2b-receptors. In contrast, CBZ does not block the inhibition elicited by adenosine-agonists of the agonist-induced increased formation of inositolphosphates in human neutrophils, which is mediated by high-affinity A2a-receptors. The specific antagonism by CBZ of A1- but not of high-affinity A2a-receptors was further supported by binding experiments using rat brain membranes. These results suggest that the paradox of CBZs antagonistic effects at adenosine-receptors might be at least partially reconciled by a selective antagonistic action of CBZ at A1 receptors but not at high-affinity A2a-receptors.

Corticotropin Peptides and Melanotropins Elevate the Level of Adenosine 3′: 5′‐Cyclic Monophosphate in Cultured Murine Brain Cells

Abstract: Cell cultures derived from mouse and rat brain and consisting mainly of astroblasts are known to respond to several hormones by increasing or decreasing their intracellular concentration of cyclic AMP. In the present study these cultures were analyzed for their susceptibility to various additional hormonal and other neuroactive compounds. Only the peptides of the corticotropin (ACTH)/melanotropin (MSH) family were found active. Their potency for elevating the intracellular level of cyclic AMP decreases in the sequence (values for the half‐maximally stimulating concentrations, EC50, in parentheses) ACTH‐(1–24) (10 nM) > α‐, β‐MSH (30 nM) > ACTH (≥ 100 nM) > γ‐MSH, ACTH−(1‐10), −(4‐10), −(4‐11) (≥0.5 μM). The lack of additivity of the maximal effects of the peptides suggests that they all act at the same receptor. The stimulation exerted by these peptides is partially suppressed by hormones known to inhibit cyclic AMP formation in that culture, i.e., noradrenaline (acting via an α‐adrenergic receptor), adenosine (acting via an A1 receptor) and somatostatin. It is concluded that the receptors for the ACTH/MSH peptides and the inhibitory hormones are located on the same cells, presumably the astroblasts. The maximal response to ACTH and α‐and β‐MSH depends strongly on the age of culture. The results are discussed in view of the facts that (1) peptides of the ACTH/MSH family affect behavior and learning in animals and (2) ACTH and α‐MSH occur in brain.

Stimulation by Bradykinin, Angiotensin II, and Carbachol of the Accumulation of Inositol Phosphates in PC-12 Pheochromocytoma Cells: Differential Effects of Lithium Ions on Inositol Mono-and Polyphosphates

Abstract: Rat PC‐12 pheochromocytoma cells respond to stimulation with bradykinin, angiotensin II, and carbachol with an increased formation of labeled inositol phosphates after preincubation of the cells with [3H]inositol. Li+ potentiates greatly the agonist‐induced increase in amount of nositol mono‐, bis‐, and trisphosphate but not the increase in amount of inositol tetrakisphosphate. Separation of the isomers of inositol trisphosphate shows that the lithium‐induced increase in amount of inositol trisphosphate is due to potentiation evoked by lithium of the accumulation of inositol‐1,3,4‐trisphosphate.

Polyploid rat glioma cells: Production, oscillations of membrane potential and response to neurohormones☆

Abstract Due to their small size, electrophysiological investigation of C6 rat glioma cells by intracellular recording with microelectrodes is very difficult. In order to facilitate such electrophysiological studies, the degree of ploidy of C6 cells was increased in three successive steps. At each step complementary drug-resistant mutants of C6 were fused and hybrid cell clones isolated in selective culture medium. Cell volume was measured by electronic cell sizing and DNA content per cell assessed by impulse cytofluorimetry. In the final polyploid line, C6-4-2, both parameters were found to be increased 6-fold in comparison with the original cell line C6. In response to β-adrenergic agonists and prostaglandin E1 the intracellular concentration of cAMP was raised in C6-4-2 cells. Stable membrane potentials could be recorded from the polyploid glioma cells much more easily than from the original C6 cells. The plasma membrane of the C6-4-2 cells could hyperpolarize spontaneously with a frequency of about 1 cycle/min. Elevation of the concentration of Ca2+ ions (but not other earth alkali ions) in the medium induced the onset of the spontaneous hyperpolarizations. The membranes of the C6-4-2 cells also hyperpolarized in response to acetylcholine, γ-aminobutyric acid (GABA) and glutamate.

A calcium antagonist for the treatment of depressive episodes: Single case reports

Preclinical studies indicate that a disturbed intracellular calcium ion homeostasis is involved in the pathophysiology of affective disorders. Therefore some calcium antagonists were investigated, especially in the treatment of the manic syndrome. In the present study the calcium antagonist nimodipine was used in 10 out-patients with single or recurrent depressive episodes. As a result the mean HAMD scores changed from 26.5 to 9.9 after the individual nimodipine administration. These single case reports suggest an effective new therapy strategy for the treatment of affective dysregulations and give rise to controlled clinical studies with calcium antagonists.

Mechanism of microglia neuroprotection: Involvement of P2X7, TNFα, and valproic acid.

Recently, we have demonstrated that ramified microglia are neuroprotective in N‐methyl‐d‐aspartate (NMDA)‐induced excitotoxicity in organotypic hippocampal slice cultures (OHSCs). The present study aimed to elucidate the underlying neuron‐glia communication mechanism. It is shown here that pretreatment of OHSC with high concentrations of adenosine 5′‐triphosphate (ATP) reduced NMDA‐induced neuronal death only in presence of microglia. Specific agonists and antagonists identified the P2X7 receptor as neuroprotective receptor which was confirmed by absence of ATP‐dependent neuroprotection in P2X7‐deficient OHSC. Microglia replenished chimeric OHSC consisting of wild‐type tissue replenished with P2X7‐deficient microglia confirmed the involvement of microglial P2X7 receptor in neuroprotection. Stimulation of P2X7 in primary microglia induced tumor necrosis factor α (TNFα) release and blocking TNFα by a neutralizing antibody in OHSC abolished neuroprotection by ATP. OHSC from TNFα‐deficient mice show increased exicitoxicity and activation of P2X7 did not rescue neuronal survival in the absence of TNFα. The neuroprotective effect of valproic acid (VPA) was strictly dependent on the presence of microglia and was mediated by upregulation of P2X7 in the cells. The present study demonstrates that microglia‐mediated neuroprotection depends on ATP‐activated purine receptor P2X7 and induction of TNFα release. This neuroprotective pathway was strengthened by VPA elucidating a novel mechanism for the neuroprotective function of VPA. GLIA 2016;64:76–89

Nerve growth factor potentiates the hormone-stimulated intracellular accumulation of inositol phosphates and Ca2+ in rat PC12 pheochromocytoma cells: comparison with the effect of epidermal growth factor

Abstract: The effects of nerve growth factor (NGF) and epidermal growth factor (EGF) on the intracellular accumulation of inositol phosphates and on cytosolic free Ca2+ concentrations were studied in rat PC12 pheochromocytoma cells. Both NGF and EGF potentiate in these cells the increase in the accumulation of inositol phosphates that is elicited by bradykinin and carbachol. A corresponding potentiation was also found for the agonist‐induced increase of cytosolic Ca2+ concentrations. The effect of NGF, but not that of EGF, is abolished when the cells are preincubated with 5′‐deoxy‐5′‐methylthioadenosine, an inhibitor of S‐adenosylhomocysteine hydrolase. These results suggest that an increased response to hormones, which act via phosphoinositide‐derived second messengers, may be important in the mechanism of action of NGF and EGF.

The agonist-stimulated accumulation of inositol phosphates is attenuated in neutrophils from male patients under chronic lithium therapy

Neutrophils from 22 patients (11 men, 11 women) under chronic lithium therapy and from 22 age- and sex-matched healthy controls were assessed for the activity of the agonist-stimulated inositol-phospholipid second messenger-producing system. [3H]inositol-labeled cells were stimulated with the chemotactic peptide formylmethionylleucylphenyl-alanin (fMLP). The fMLP-evoked increase in the accumulation of [3H]inositol phosphates was significantly attenuated in neutrophils from chronically lithium-treated male but not female patients. Furthermore, the fMLP-stimulated accumulation of inositol phosphates was attenuated in neutrophils from male volunteers, when the labeling of the cells with [3H]inositol was performed in the presence of 1mM Li (4 hr, 37 degrees C). However, the presence of lithium ions during the labeling did not further reduce the already diminished response of neutrophils from patients under lithium therapy. These results suggest that lithium treatment induces an inhibition of the agonist-evoked breakdown of inositol phospholipids in human cells, as already shown for rat brain slices.

Interpersonelle Psychotherapie bei stationär behandelten, depressiven Patienten

Zusammenfassung. Theoretischer Hintergrund: Eine erfolgreiche Depressionsbehandlung beinhaltet neben einer Remission depressiver Symptome, die Verbesserung der psychosozialen Leistungsfahigkeit. Fragestellung: Fuhrt eine stationare Behandlung mit Interpersoneller Psychotherapie in Kombination mit Medikation zu ausgepragteren psychosozialen und interpersonellen Verbesserungen im Vergleich zu einer Standardbehandlung? Methode: 105 depressive Patienten, die auf „Interpersonelle Psychotherapie plus Pharmakotherapie” oder „Pharmakotherapie plus arztliche Kurzgesprache” randomisiert wurden, wurden nach 5 Wochen stationarer Behandlung sowie 3 und 12 Monate nach Entlassung untersucht. Ergebnisse: In der klinischen Beurteilung der allgemeinen psychosozialen Leistungsfahigkeit erwies sich die Kombinationsbehandlung der Standardbehandlung signifikant uberlegen, was allerdings uber eine Verbesserung der Depressivitat zu erklaren war. In der Selbstbeurteilung der sozialen und interpersonellen Anpassung zeigten sich ke...