Laurent Tritschler

The Postischemic Environment Differentially Impacts Teratoma or Tumor Formation After Transplantation of Human Embryonic Stem Cell-Derived Neural Progenitors

Background and Purpose— Risk of tumorigenesis is a major obstacle to human embryonic and induced pluripotent stem cell therapy. Likely linked to the stage of differentiation of the cells at the time of implantation, formation of teratoma/tumors can also be influenced by factors released by the host tissue. We have analyzed the relative effects of the stage of differentiation and the postischemic environment on the formation of adverse structures by transplanted human embryonic stem cell-derived neural progenitors. Methods— Four differentiation stages were identified on the basis of quantitative polymerase chain reaction expression of pluripotency, proliferation, and differentiation markers. Neural progenitors were transplanted at these 4 stages into rats with no, small, or large middle cerebral artery occlusion lesions. The fate of each transplant was compared with their pretransplantation status 1 to 4 months posttransplantation. Results— The influence of the postischemic environment was limited to graft survival and occurrence of nonneuroectodermal structures after transplantation of very immature neural progenitors. Both effects were lost with differentiation. We identified a particular stage of differentiation characterized in vitro by a rebound of proliferative activity that produced highly proliferative grafts susceptible to threaten surrounding host tissues. Conclusion— The effects of the ischemic environment on the formation of teratoma by transplanted human embryonic stem cell-derived neural progenitors are limited to early differentiation stages that will likely not be used for stem cell therapy. In contrast, hyperproliferation observed at later stages of differentiation corresponds to an intrinsic activity that should be monitored to avoid tumorigenesis.

Learning and memory impairments in a neuroendocrine mouse model of anxiety/depression.

Cognitive disturbances are often reported as serious incapacitating symptoms by patients suffering from major depressive disorders (MDDs). Such deficits have been observed in various animal models based on environmental stress. Here, we performed a complete characterization of cognitive functions in a neuroendocrine mouse model of depression based on a chronic (4 weeks) corticosterone administration (CORT). Cognitive performances were assessed using behavioral tests measuring episodic (novel object recognition test, NORT), associative (one-trial contextual fear conditioning, CFC), and visuo-spatial (Morris water maze, MWM; Barnes maze, BM) learning/memory. Altered emotional phenotype after chronic corticosterone treatment was confirmed in mice using tests predictive of anxiety or depression-related behaviors. In the NORT, CORT-treated mice showed a decrease in time exploring the novel object during the test session and a lower discrimination index compared to control mice, characteristic of recognition memory impairment. Associative memory was also impaired, as observed with a decrease in freezing duration in CORT-treated mice in the CFC, thus pointing out the cognitive alterations in this model. In the MWM and in the BM, spatial learning performance but also short-term spatial memory were altered in CORT-treated mice. In the MWM, unlike control animals, CORT-treated animals failed to learn a new location during the reversal phase, suggesting a loss of cognitive flexibility. Finally, in the BM, the lack of preference for the target quadrant during the recall probe trial in animals receiving corticosterone regimen demonstrates that long-term retention was also affected in this paradigm. Taken together, our results highlight that CORT-induced anxio-depressive-like phenotype is associated with a cognitive deficit affecting all aspects of memory tested.

Combined 192 IgG-saporin and 5,7-dihydroxytryptamine lesions in the male rat brain: A neurochemical and behavioral study

In a previous experiment [Eur J Neurosci 12 (2000) 79], combined intracerebroventricular injections of 5,7-dihydroxytryptamine (5,7-DHT; 150 microg) and 192 IgG-saporin (2 microg) in female rats produced working memory impairments, which neither single lesion induced. In the present experiment, we report on an identical approach in male rats. Behavioral variables were locomotor activity, T-maze alternation, beam-walking, Morris water-maze (working and reference memory) and radial-maze performances. 192 IgG-saporin reduced cholinergic markers in the frontoparietal cortex and the hippocampus. 5,7-DHT lesions reduced serotonergic markers in the cortex, hippocampus and striatum. Cholinergic lesions induced motor deficits, hyperactivity and reduced T-maze alternation, but had no other effect. Serotonergic lesions only produced hyperactivity and reduced T-maze alternation. Beside the deficits due to cholinergic lesions, rats with combined lesions also showed impaired radial-maze performances. We confirm that 192 IgG-saporin and 5,7-DHT injections can be combined to produce concomitant damage to cholinergic and serotonergic neurons in the brain. In female rats, this technique enabled to show that interactions between serotonergic and basal forebrain cholinergic mechanisms play an important role in cognitive functions. The results of the present experiment in male rats are not as clear-cut, although they are not in obvious contradiction with our previous results in females.

Nrf2-signaling and BDNF: A new target for the antidepressant-like activity of chronic fluoxetine treatment in a mouse model of anxiety/depression

Several studies have shown that Nrf2, a major redox-sensitive transcription factor involved in the cellular defense against oxidative stress, increases susceptibility to depressive-like behavior. However, little is known about the influence of antidepressant drugs on Nrf2 signaling and expression of its target genes (GCLC, NQO1, HO-1) in the brain. We found that chronic treatment of a mouse model of anxiety/depression (CORT model) with a selective serotonin reuptake inhibitor (SSRI, fluoxetine, 18mg/kg/day) reversed CORT-induced anxiety/depression-like behavior in mice. Chronic fluoxetine treatment restored CORT-induced decreases in Nrf2 protein levels and its target genes in the cortex and hippocampus. Furthermore, we found that chronic fluoxetine also increased brain derived neurotrophic factor (BDNF) protein levels in cortex and hippocampus of CORT-treated Nrf2 knockout mice (KO, Nrf2(-/-)). Taken together, these data suggest that Nrf2 signaling contributes to fluoxetine-induced neuroprotection via an unexpected mechanism involving 5-HT transporter SERT blockade, and not through enhancement of BDNF expression.

Distinct circuits underlie the effects of 5-HT1b receptors on aggression and impulsivity

Impulsive and aggressive behaviors are both modulated by serotonergic signaling, specifically through the serotonin 1B receptor (5-HT1BR). 5-HT1BR knockout mice show increased aggression and impulsivity, and 5-HT1BR polymorphisms are associated with aggression and drug addiction in humans. To dissect the mechanisms by which the 5-HT1BR affects these phenotypes, we developed a mouse model to spatially and temporally regulate 5-HT1BR expression. Our results demonstrate that forebrain 5-HT1B heteroreceptors expressed during an early postnatal period contribute to the development of the neural systems underlying adult aggression. However, distinct heteroreceptors acting during adulthood are involved in mediating impulsivity. Correlating with the impulsivity, dopamine in the nucleus accumbens is elevated in the absence of 5-HT1BRs and normalized following adult rescue of the receptor. Overall, these data show that while adolescent expression of 5-HT1BRs influences aggressive behavior, a distinct set of 5-HT1B receptors modulates impulsive behavior during adulthood.

Ketamine treatment involves medial prefrontal cortex serotonin to induce a rapid antidepressant-like activity in BALB/cJ mice.

ABSTRACT Unlike classic serotonergic antidepressant drugs, ketamine, an NMDA receptor antagonist, exhibits a rapid and persistent antidepressant (AD) activity, at sub‐anaesthetic doses in treatment‐resistant depressed patients and in preclinical studies in rodents. The mechanisms mediating this activity are unclear. Here, we assessed the role of the brain serotonergic system in the AD‐like activity of an acute sub‐anaesthetic ketamine dose. We compared ketamine and fluoxetine responses in several behavioral tests currently used to predict anxiolytic/antidepressant‐like potential in rodents. We also measured their effects on extracellular serotonin levels [5‐HT]ext in the medial prefrontal cortex (mPFCx) and brainstem dorsal raphe nucleus (DRN), a serotonergic nucleus involved in emotional behavior, and on 5‐HT cell firing in the DRN in highly anxious BALB/cJ mice. Ketamine (10 mg/kg i.p.) had no anxiolytic‐like effect, but displayed a long lasting AD‐like activity, i.e., 24 h post‐administration, compared to fluoxetine (18 mg/kg i.p.). Ketamine (144%) and fluoxetine (171%) increased mPFCx [5‐HT]ext compared to vehicle. Ketamine‐induced AD‐like effect was abolished by a tryptophan hydroxylase inhibitor, para‐chlorophenylalanine (PCPA) pointing out the role of the 5‐HT system in its behavioral activity. Interestingly, increase in cortical [5‐HT]ext following intra‐mPFCx ketamine bilateral injection (0.25 &mgr;g/side) was correlated with its AD‐like activity as measured on swimming duration in the FST in the same mice. Furthermore, pre‐treatment with a selective AMPA receptor antagonist (intra‐DRN NBQX) blunted the effects of intra‐mPFCx ketamine on both the swimming duration in the FST and mPFCx [5‐HT]ext suggesting that the AD‐like activity of ketamine required activation of DRN AMPA receptors and recruited the prefrontal cortex/brainstem DRN neural circuit in BALB/c mice. These results confirm a key role of cortical 5‐HT release in ketamines AD‐like activity following the blockade of glutamatergic NMDA receptors. Tight interactions between mPFCx glutamatergic and serotonergic systems may explain the differences in this activity between ketamine and fluoxetine in vivo. This article is part of the Special Issue entitled ‘Ionotropic glutamate receptors’. HIGHLIGHTSUnlike fluoxetine, ketamine induced persistent antidepressant‐like effect in mice.Ketamine increase cortical, but not [5‐HT]ext in the dorsal raphe nucleus.Increases in cortical 5‐HT is required for ketamines antidepressant‐like activity.Activation of DRN AMPA receptors and of the prefrontal cortex/brainstem DRN neural circuit are required for ketamines antidepressant‐like activity in BALB/c mice.

Vortioxetine for the treatment of major depressive disorder

Vortioxetine (Brintellix®, 1-[2-(2,4-dimethylphenyl-sulfanyl)-phenyl]-piperazine) is a multimodal antidepressant targeting the 5-HT1A, 5-HT1B, 5-HT1D, 5-HT3, 5-HT7 receptors and the serotonin (5-HT) transporter (5-HTT). Vortioxetine administration induces antidepressant- and anxiolytic-like effects, and can enhance cognitive performance in rodents. Several clinical trials have reported the efficiency and a satisfactory tolerability of vortioxetine treatment in depressed patients. Remarkably, vortioxetine has a specific positive impact on cognitive symptoms in depressed patients. Overall, vortioxetine is an efficacious antidepressant drug for the treatment of patients with a major depressive episode and has a unique mechanism of action offering a new therapeutic option.

A Lack of Serotonin 1B Autoreceptors Results in Decreased Anxiety and Depression-Related Behaviors.

The effects of serotonin (5-HT) on anxiety and depression are mediated by a number of 5-HT receptors, including autoreceptors that act to inhibit 5-HT release. While the majority of anxiety and depression-related research has focused on the 5-HT1A receptor, the 5-HT1B receptor has a lesser known role in modulating emotional behavior. 5-HT1B receptors are inhibitory GPCRs located on the presynaptic terminal of both serotonin and non-serotonin neurons, where they act to inhibit neurotransmitter release. The autoreceptor population located on the axon terminals of 5-HT neurons is a difficult population to study due to their diffuse localization throughout the brain that overlaps with 5-HT1B heteroreceptors (receptors located on non-serotonergic neurons). In order to study the contribution of 5-HT1B autoreceptors to anxiety and depression-related behaviors, we developed a genetic mouse model that allows for selective ablation of 5-HT1B autoreceptors. Mice lacking 5-HT1B autoreceptors displayed the expected increases in extracellular serotonin levels in the ventral hippocampus following administration of a selective serotonin reuptake inhibitor. In behavioral studies, they displayed decreased anxiety-like behavior in the open field and antidepressant-like effects in the forced swim and sucrose preference tests. These results suggest that strategies aimed at blocking 5-HT1B autoreceptors may be useful for the treatment of anxiety and depression.

Pro-inflammatory mediators and apoptosis correlate to rt-PA response in a novel mouse model of thromboembolic stroke.

Background A recent study suggests that patients with persistent occlusion of the middle cerebral artery (MCA) following treatment with recombinant tissue plasminogen activator (rt-PA) have better outcomes than patients with MCA occlusion not receiving rt-PA. We performed a study to elucidate possible mechanisms of this finding in a new model of thromboembolic stroke closely mimicking human pathophysiology. Methods Thromboembolic stroke was induced by local injection of thrombin directly into the right MCA of C57 black/6J mice. Rt-PA was administered 20 and 40 min after clot formation. The efficiency of rt-PA to induce thrombolysis was measured by laser Doppler. After 24 h, all animals were euthanized and interleukin (IL)-6, tumor necrosis factor-alpha (TNF-α), matrix metalloproteinase (MMP)-9, Caspase-3, hsp 32 and hsp 70 protein levels were investigated by immunofluorescence. Presence of hemorrhage was verified and infarct volume was measured using histology. Results Thrombin injection resulted in clot formation giving rise to cortical brain infarction. Early rt-PA treatment starting at 20 min after the clot formation resulted in 100% recanalization. However, rt-PA-induced thrombolysis dissolved the clot in only 38% of the animals when administered 40 min after clot formation. Protein levels of IL-6, TNF-α, MMP-9, Caspase-3, hsp 32 and hsp 70 were increased after MCAO, whereas treatment with rt-PA attenuated the expressions of inflammatory markers in those animals where the thrombolysis was successful. In addition, the infarct size was significantly reduced with rt-PA treatment compared to non-treated MCAO, regardless of whether MCA thrombolysis was successful. Conclusions The present study demonstrates a clear correlation of the protein expression of inflammatory mediators, apoptosis and stress genes with the recanalization data after rt-PA treatment. In this model rt-PA treatment decreases the infarct size regardless of whether vessel recanalization is successful.

A functional subdivision of the circadian clock is revealed by differential effects of melatonin administration

The biological clock of the suprachiasmatic nuclei drives numerous physiological and behavioural circadian rhythms. In this study, we addressed the question as to whether different components of the clock may control separately various circadian functions. Using the rat transpineal microdialysis tool, we analysed the effect of clock perturbation by exogenous melatonin injection on two hormonal clock outputs: pineal melatonin and adrenal corticosterone secretions. As already reported, a single melatonin injection at the light/dark transition induces a marked increase in the endogenous pineal melatonin peak for the two following days. In the same animals, by contrast, the amplitude of the corticosterone rhythm was not altered following melatonin injection. These data show that the melatonin injection does not display an overall effect on the circadian clock, but rather influences a subpopulation of melatonin-sensitive neurons involved, among other functions, in the circadian control of the pineal pathway.