No abstract

Serotonin synthesis in mammals is initiated by 2 distinct tryptophan hydroxylases (TPH), TPH1 and TPH2. By genetically ablating TPH2, we created mice (Tph2 ؊/؊ ) that lack serotonin in the central nervous system. Surprisingly, these mice can be born and survive until adulthood. However, depletion of serotonin signaling in the brain leads to growth retardation and 50% lethality in the first 4 weeks of postnatal life. Telemetric monitoring revealed more extended daytime sleep, suppressed respiration, altered body temperature control, and decreased blood pressure (BP) and heart rate (HR) during nighttime in Tph2 ؊/؊ mice. Moreover, Tph2 ؊/؊ females, despite being fertile and producing milk, exhibit impaired maternal care leading to poor survival of their pups. These data confirm that the majority of central serotonin is generated by TPH2. TPH2-derived serotonin is involved in the regulation of behavior and autonomic pathways but is not essential for adult life.growth retardation ͉ maternal care ͉ respiration ͉ serotonin ͉ sleep S erotonin (5-hydroxytryptamine, 5-HT) is an extracellular signaling molecule with a multitude of functions in the central nervous system (CNS) and in the periphery. 5-HT effects are conveyed by at least 13 receptors classified in 7 families, 5-HT1 to 5-HT7. Serotonin synthesis from tryptophan is initiated by the enzyme tryptophan hydroxylase (TPH) generating 5-hydroxytryptophan followed by aromatic amino acid decarboxylase (AADC), which produces 5-HT. We have recently discovered that 2 TPH isoenzymes exist in all vertebrates, TPH1 and TPH2, encoded by 2 distinct genes (1, 2). Tph1 is mainly expressed in the gut, generating serotonin that is distributed into the whole body by thrombocytes, and in the pineal gland, where the resulting 5-HT is metabolized to melatonin. The Tph1-deficient mice generated by us (2) and others (3, 4) revealed that 95% of peripheral 5-HT is produced by TPH1. They also revealed that 5-HT in platelets and other peripheral cells is involved in such diverse processes as thrombosis (5), liver regeneration (6), hepatitis (7), colon cancer (8), mammary gland plasticity (9), pulmonary hypertension (10), and bone formation (11). TPH2, on the other hand, is responsible for the synthesis of serotonin in the raphé nuclei of the brainstem, from where all central serotonergic projections originate (12). Accordingly, polymorphisms and functional mutations in the human and mouse genes for this enzyme have been linked to neurological and behavioral abnormalities (13)(14)(15)(16).In this study, we generated mice lacking TPH2 by gene targeting and analyzed the physiological consequences resulting from a lack of brain serotonin. Results and DiscussionGeneration and Basic Characteristics of Tph2-Deficient Mice. Tph2-deficient (Tph2 Ϫ/Ϫ ) mice were generated by deleting the coding sequence in exons 1 and 2 (supporting information (SI) Fig. S1 A and B). In the resulting Tph2 Ϫ/Ϫ mice, no Tph2 mRNA could be found by RT-PCR (Fig. S1C) and in situ hybridization in the brain (Fig. 1A). Immu...

Altered glucocorticoid receptor (GR) signaling is a postulated mechanism for the pathogenesis of major depression. To mimic the human situation of altered GR function claimed for depression, we generated mouse strains that underexpress or overexpress GR, but maintain the regulatory genetic context controlling the GR gene. To achieve this goal, we used the following: (1) GR-heterozygous mutant mice (GR ϩ/Ϫ ) with a 50% GR gene dose reduction, and (2) mice overexpressing GR by a yeast artificial chromosome resulting in a twofold gene dose elevation. GR ϩ/Ϫ mice exhibit normal baseline behaviors but demonstrate increased helplessness after stress exposure, a behavioral correlate of depression in mice. Similar to depressed patients, GR ϩ/Ϫ mice have a disinhibited hypothalamic-pituitary-adrenal (HPA) system and a pathological dexamethasone/corticotropin-releasing hormone test. Thus, they represent a murine depression model with good face and construct validity. Overexpression of GR in mice evokes reduced helplessness after stress exposure, and an enhanced HPA system feedback regulation. Therefore, they may represent a model for a stress-resistant strain. These mouse models can now be used to study biological changes underlying the pathogenesis of depressive disorders. As a first potential molecular correlate for such changes, we identified a downregulation of BDNF protein content in the hippocampus of GR ϩ/Ϫ mice, which is in agreement with the so-called neurotrophin hypothesis of depression.

Graphical abstractHighlights► The distribution of GABAA receptor subunits is highly heterogeneous. ► The distribution of mRNAs corresponds to that of proteins. ► The distribution in the mouse correlates largely to that in rats although there are distinct differences.

After mild ischemic insults, many neurons undergo delayed neuronal death. Aberrant activation of the cell cycle machinery is thought to contribute to apoptosis in various conditions including ischemia. We demonstrate that loss of endogenous cyclin-dependent kinase (Cdk) inhibitor p16INK4a is an early and reliable indicator of delayed neuronal death in striatal neurons after mild cerebral ischemia in vivo. Loss of p27 Kip1 , another Cdk inhibitor, precedes cell death in neocortical neurons subjected to oxygen-glucose deprivation in vitro. The loss of Cdk inhibitors is followed by upregulation of cyclin D1, activation of Cdk2, and subsequent cytoskeletal disintegration. Most neurons undergo cell death before entering S-phase, albeit a small number (ϳ1%) do progress to the S-phase before their death. Treatment with Cdk inhibitors significantly reduces cell death in vitro. These results show that alteration of cell cycle regulatory mechanisms is a prelude to delayed neuronal death in focal cerebral ischemia and that pharmacological interventions aimed at neuroprotection may be usefully directed at cell cycle regulatory mechanisms.

Several lines of clinical and experimental evidence suggest an important role of the renin-angiotensin system in ischemic brain injury although the cellular regulation of the angiotensin AT1 and AT2 receptors and their potential relevance in this condition have not yet been clearly defined. We first assessed the regulation of brain AT1 and AT2 receptors in response to transient unilateral medial cerebral artery occlusion in rats by real-time RT-PCR, Western blot, and immunofluorescence labeling. AT2 receptors in the peri-infarct zone were significantly upregulated 2 days after transient focal cerebral ischemia. Increased AT2 receptors, which were abundantly distributed in a large number of brain regions adjacent to the infarct area including cerebral frontal cortex, piriform cortex, striatum, and hippocampus, were exclusively expressed in neurons. By contrast, AT1 receptors, which remained unaltered, were mainly expressed in astrocytes. In neurons of ischemic striatum, increased AT2 receptors were associated with intense neurite outgrowth. Blockade of central AT2 receptors with PD123177 abolished the neuroprotective effects of central AT1 receptor blockade with irbesartan on infarct size and neurological outcome. In primary cortical neurons, stimulation of AT2 receptors supported neuronal survival and neurite outgrowth. Our data indicate that cerebral AT2 receptors exert neuroprotective actions in response to ischemia-induced neuronal injury, possibly by supporting neuronal survival and neurite outgrowth in peri-ischemic brain areas.

No abstract

Statins [3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors] exert cholesterol-independent pleiotropic effects that include anti-thrombotic, anti-inflammatory, and antioxidative properties. Here, we examined direct protective effects of atorvastatin on neurones in different cell damage models in vitro. Primary cortical neurones were pre-treated with atorvastatin and then exposed to (i) glutamate, (ii) oxygenglucose deprivation or (iii) several apoptosis-inducing compounds. Atorvastatin significantly protected from glutamate-induced excitotoxicity as evidenced by propidium iodide staining, nuclear morphology, release of lactate dehydrogenase, and mitochondrial tetrazolium metabolism, but not from oxygen-glucose deprivation or apoptotic cell death. This antiexcitototoxic effect was evident with 2-4 days pre-treatment but not with daily administration or shorter-term pre-treatment. The protective properties occurred independently of 3-hydroxy-3-methylglutaryl-CoA reductase inhibition because co-treatment with mevalonate or other isoprenoids did not reverse or attenuate neuroprotection. Atorvastatin attenuated the glutamate-induced increase of intracellular calcium, which was associated with a modulation of NMDA receptor function. Taken together, atorvastatin exerts specific anti-excitotoxic effects independent of 3-hydroxy-3-methylglutaryl-CoA reductase inhibition, which has potential therapeutic implications.