Effect of Hippocampal Overexpression of Dopamine Neurotrophic Factor (CDNF) on Behavior of Mice with Genetic Predisposition to Depressive-Like Behavior.

Cerebral dopamine neurotrophic factor (CDNF) is a promising agent for Parkinson's disease treatment. However, its role in regulation of non-motor behavior including various psychopathologies remains unclear. In this regard, the aim of the present work was to study effect of CDNF overexpression in hippocampus on behavior of the ASC mice (Antidepressant Sensitive Cataleptics) with genetic predisposition to depressive-like behavior. CDNF overexpression in the mouse hippocampal neurons was induced using an adeno-associated viral vector. Four weeks after stereotaxic injection of the AAV-CDNF construct into the dorsal hippocampus home cage activity, exploratory, anxious and depressive-like types of behavior, as well as spatial and associative learning were assessed. We found significant improvements in the dynamics of spatial learning in the Morris water maze in the CDNF-overexpressing animals. At the same time, no effect of CDNF was found on other types of behavior under study. Behavior of the experimental animals under home cage conditions did not differ from that in the control group, except for the decrease in the total amount of food eaten and slight increase in the number of sleep episodes during the light phase of the day. In the present study we also attempted to determine molecular basis for the above-mentioned changes through assessment of the gene expression pattern. We did not find significant changes in the mRNA level of key kinases genes involved in neuroplasticity and neuronal survival, as well as genes encoding receptors for the main neurotransmitter systems. However, the CDNF-overexpressing animals showed increased level of the spliced Xbp indicating activation of the Ire1α/Xbp-1 pathway traditionally associated with ER stress. Immunohistochemical analysis showed that CDNF was co-localized with the ER marker calreticulin. Thus, the effects of endogenous CDNF on behavior that we have found could be mediated by a specific molecular cascade, which emphasizes its difference from the classical neurotrophic factors.

Amisulpride Decreases Tau Protein Hyperphosphorylation in the Brain of OXYS Rats.

AIM: In this study, OXYS rats of three ages (1, 3, and 6 months), a proven model of Alzheimer's disease (AD), at various stages of disease progression were used to thoroughly study the effects of amisulpride on behavior and tau protein phosphorylation. BACKGROUND: With the growing number of patients with AD, the problem of finding a cure is very acute. Neurodegeneration in AD has various causes, one of which is hyperphosphorylation of tau protein. OBJECTIVE: This study aimed to investigate whether amisulpride would affect pathological tau phosphorylation in AD. METHODS: We assessed the influence of chronic administration of amisulpride (3 weeks, 3 mg/kg per day, intraperitoneally)-a 5-HT7 receptor inverse agonist-on behavior and tau hyperphosphorylation in OXYS rats (at ages of 1, 3, and 6 months). RESULTS: Chronic administration of amisulpride dramatically decreased tau phosphorylation in the frontal cortex and hippocampus of 3-month-old OXYS rats. Additionally, in 1- and 3-month-old rats' hippocampi, amisulpride diminished the mRNA level of the Cdk5 gene encoding one of the main tau kinases involved in the 5-HT7 receptor-induced effect on tau phosphorylation. CONCLUSION: Thus, We found that chronic administration of amisulpride could reduce pathological tau hyperphosphorylation while reducing anxiety. We propose amisulpride to have therapeutic potential against AD and that it can be the most effective in the early stages of the disease.

On the role of serotonin 5-HT1A receptor in autistic-like behavior: сross talk of 5-HT and BDNF systems.

Autism spectrum disorders (ASDs) are some of the most common neurodevelopmental disorders; however, the mechanisms underlying ASDs are still poorly understood. Serotonin (5-HT) and brain-derived neurotrophic factor (BDNF) are known as key players in brain and behavioral plasticity and interact with each other. 5-HT1A receptor is a principal regulator of the brain 5-HT system, which modulates normal and pathological behavior. Here we investigated effects of adeno-associated-virus-based 5-HT1A receptor overexpression in the hippocampus of BTBR mice (which are a model of autism) on various types of behavior and on the expression of 5-HT7 receptor, proBDNF, mature BDNF, and BDNF receptors (TrkB and p75NTR). The 5-HT1A receptor overexpression in BTBR mice reduced stereotyped behavior in the marble-burying test and extended the time spent in the center in the open field test. Meanwhile, this overexpression failed to affect social behavior in the three-chambered test, immobility time in the tail suspension test, locomotor activity in the open field test, and associative learning within the "operant wall" paradigm. The 5-HT1A receptor overexpression in the hippocampus raised hippocampal 5-HT7 receptor mRNA and protein levels. Additionally, the 5-HT1A receptor overexpression lowered both mRNA and protein levels of TrkB receptor but failed to affect proBDNF, mature BDNF, and p75NTR receptor expression in the hippocampus of BTBR mice. Thus, obtained results suggest the involvement of the 5-HT and BDNF systems' interaction mediated by 5-HT1A and TrkB receptors in the mechanisms underlying autistic-like behavior in BTBR mice.

Effects of the Cc2d1a/Freud-1 Knockdown in the Hippocampus of BTBR Mice on the Autistic-Like Behavior, Expression of Serotonin 5-HT1A and D2 Dopamine Receptors, and CREB and NF-kB Intracellular Signaling.

The mechanisms of autism are of extreme interest due to the high prevalence of this disorder in the human population. In this regard, special attention is given to the transcription factor Freud-1 (encoded by the Cc2d1a gene), which regulates numerous intracellular signaling pathways and acts as a silencer for 5-HT1A serotonin and D2 dopamine receptors. Disruption of the Freud-1 functions leads to the development of various psychopathologies. In this study, we found an increase in the expression of the Cc2d1a/Freud-1 gene in the hippocampus of BTBR mice (model of autistic-like behavior) in comparison with C57Bl/6J mice and examined how restoration of the Cc2d1a/Freud-1 expression in the hippocampus of BTBR mice affects their behavior, expression of 5-HT1A serotonin and D2 dopamine receptors, and CREB and NF-κB intracellular signaling pathways in these animals. Five weeks after administration of the adeno-associated viral vector (AAV) carrying the pAAV_H1-2_shRNA-Freud-1_Syn_EGFP plasmid encoding a small hairpin RNA (shRNA) that suppressed expression of the Cc2d1a/Freud-1 gene, we observed an elevation in the anxiety levels, as well as the increase in the escape latency and path length to the platform in the Morris water maze test, which was probably associated with a strengthening of the active stress avoidance strategy. However, the Cc2d1a/Freud-1 knockdown did not affect the spatial memory and phosphorylation of the CREB transcription factor, although such effect was found in C57Bl/6J mice in our previous study. These results suggest the impairments in the CREB-dependent effector pathway in BTBR mice, which may play an important role in the development of the autistic-like phenotype. The knockdown of Cc2d1a/Freud-1 in the hippocampus of BTBR mice did not affect expression of the 5-HT1A serotonin and D2 dopamine receptors and key NF-κB signaling genes (Nfkb1 and Rela). Our data suggest that the transcription factor Freud-1 plays a significant role in the pathogenesis of anxiety and active stress avoidance in autism.

Serotonin 5-HT7 receptor overexpression in the raphe nuclei area produces antidepressive effect and affects brain serotonin system in male mice.

Heterodimerization between 5-HT7 and 5-HT1A receptors seems to play an important role in the mechanism of depression and antidepressant drug action. It was suggested that the shift of the ratio between 5-HT1A /5-HT7 hetero- and 5-HT1A /5-HT1A homodimers in presynaptic neurons toward 5-HT1A /5-HT1A homodimers is one of the reasons of depression. Consequently, the artificial elevation of 5-HT7 receptor number in presynaptic terminals might restore physiological homo-/heterodimer ratio resulting in antidepressive effect. Here we showed that adeno-associated virus (AAV)-based 5-HT7 receptor overexpression in the midbrain raphe nuclei area produced antidepressive effect in male mice of both C57Bl/6J and genetically predisposed to depressive-like behavior ASC (antidepressant sensitive cataleptics) strains. These changes were accompanied by the elevation of 5-HT7 receptor mRNA level in the frontal cortex of C57Bl/6J and its reduction in the hippocampus of ASC mice. The presence of engineered 5-HT7 receptor in the midbrain of both mouse strains was further demonstrated. Importantly that 5-HT7 receptor overexpression resulted in the reduction of 5-HT1A receptor level in the membrane protein fraction from the midbrain samples of C57Bl/6J, but not ASC, mice. 5-HT7 receptor overexpression caused an increase of 5-HIAA/5-HT ratio in the midbrain and the frontal cortex of C57Bl/6J and in all investigated brain structures of ASC mice. Thus, 5-HT7 receptor overexpression in the raphe nuclei area affects brain 5-HT system and causes antidepressive effect both in C57Bl/6J and in "depressive" ASC male mice. Obtained results indicate the involvement of 5-HT7 receptor in the mechanisms underlying depressive behavior.

Effects of a Cc2d1a/Freud-1 Knockdown in the Hippocampus on Behavior, the Serotonin System, and BDNF.

The serotonin 5-HT1A receptor is one of the most abundant and widely distributed brain serotonin (5-HT) receptors that play a major role in the modulation of emotions and behavior. The 5-HT1A receptor gene (Htr1a) is under the control of transcription factor Freud-1 (also known as Cc2d1a/Freud-1). Here, using adeno-associated virus (AAV) constructs in vivo, we investigated effects of a Cc2d1a/Freud-1 knockdown in the hippocampus of C57BL/6J mice on behavior, the brain 5-HT system, and brain-derived neurotrophic factor (BDNF). AAV particles carrying the pAAV_H1-2_shRNA-Freud-1_Syn_EGFP plasmid encoding a short-hairpin RNA targeting mouse Cc2d1a/Freud-1 mRNA had an antidepressant effect in the forced swim test 5 weeks after virus injection. The knockdown impaired spatiotemporal memory as assessed in the Morris water maze. pAAV_H1-2_shRNA-Freud-1_Syn_EGFP decreased Cc2d1a/Freud-1 mRNA and protein levels. Furthermore, the Cc2d1a/Freud-1 knockdown upregulated 5-HT and its metabolite 5-hydroxyindoleacetic acid but not their ratio. The Cc2d1a/Freud-1 knockdown failed to increase mRNA and protein levels of Htr1a but diminished a 5-HT1A receptor functional response. Meanwhile, the Cc2d1a/Freud-1 knockdown reduced Creb mRNA expression and CREB phosphorylation and upregulated cFos mRNA. The knockdown enhanced the expression of a BDNF precursor (proBDNF protein), which is known to play a crucial part in neuroplasticity. Our data indicate that transcription factor Cc2d1a/Freud-1 is implicated in the pathogenesis of depressive disorders not only via the 5-HT1A receptor and transcription factor CREB but also through an influence on BDNF.

Cerebral Dopamine Neurotrophic Factor (CDNF): Structure, Functions, and Therapeutic Potential.

The cerebral dopamine neurotrophic factor (CDNF) together with the mesencephalic astrocyte-derived neurotrophic factor (MANF) form a unique family of neurotrophic factors (NTFs) structurally and functionally different from other proteins with neurotrophic activity. CDNF has no receptors on the cell membrane, is localized mainly in the cavity of endoplasmic reticulum (ER), and its primary function is to regulate ER stress. In addition, CDNF is able to suppress inflammation and apoptosis. Due to its functions, CDNF has demonstrated outstanding protective and restorative properties in various models of neuropathology associated with ER stress, including Parkinson's disease (PD). That is why CDNF already passed clinical trials in patients with PD. However, despite the name, CDNF functions extend far beyond the dopamine system in the brain. In particular, there are data on participation of CDNF in the maturation and maintenance of other neurotransmitter systems, regulation of the processes of neuroplasticity and non-motor behavior. In the present review, we discuss the features of CDNF structure and functions, its protective and regenerative properties.

Genetic Background Underlying 5-HT1A Receptor Functioning Affects the Response to Fluoxetine.

The influence of genetic background on sensitivity to drugs represents a topical problem of personalized medicine. Here, we investigated the effect of chronic (20 mg/kg, 14 days, i.p.) antidepressant fluoxetine treatment on recombinant B6-M76C mice, differed from control B6-M76B mice by CBA-derived 102.73-110.56 Mbp fragment of chromosome 13 and characterized by altered sensitivity of 5-HT1A receptors to chronic 8-OH-DPAT administration and higher 5-HT1A receptor mRNA levels in the frontal cortex and hippocampus. Significant changes in the effects of fluoxetine treatment on behavior and brain 5-HT system in recombinant B6-M76C mice were revealed. In contrast to B6-M76B mice, in B6-M76C mice, fluoxetine produced pro-depressive effects, assessed in a forced swim test. Fluoxetine decreased 5-HT1A receptor mRNA levels in the cortex and hippocampus, reduced 5-HT1A receptor protein levels and increased receptor silencer Freud-1 protein levels in the hippocampus of B6-M76C mice. Fluoxetine increased mRNA levels of the gene encoding key enzyme for 5-HT synthesis in the brain, tryptophan hydroxylase-2, but decreased tryptophan hydroxylase-2 protein levels in the midbrain of B6-M76B mice. These changes were accompanied by increased expression of the 5-HT transporter gene. Fluoxetine reduced 5-HT and 5-HIAA levels in cortex, hippocampus and midbrain of B6-M76B and in cortex and midbrain of B6-M76C; mice. These data demonstrate that changes in genetic background may have a dramatic effect on sensitivity to classic antidepressants from the Selective Serotonin Reuptake Inhibitors family. Additionally, the results provide new evidence confirming our idea on the disrupted functioning of 5-HT1A autoreceptors in the brains of B6-M76C mice, suggesting these mice as a model of antidepressant resistance.

The Chronic Treatment With 5-HT2A Receptor Agonists Affects the Behavior and the BDNF System in Mice.

Serotonin 5-HT2A receptors and the brain-derived neurotrophic factor (BDNF) are involved in the pathophysiology and treatment of many psychiatric diseases. However, the interaction between 5-HT2A and BDNF is still poorly understood. In the present paper, the effects of chronic treatment with mixed 5-HT2A/2C receptor agonist DOI, highly selective 5-HT2A agonists TCB-2 and 25CN-NBOH on behavior and the BDNF system have been investigated. Chronic treatment of males of C57Bl/6 mice with DOI, TCB-2 and 25CN-NBOH (1 mg/kg, i.p., 14 days) resulted in desensitization of 5-HT2A receptors. Treatment with 25CN-NBOH significantly increased startle amplitude. At the same time all used drugs failed to affect anxiety, exploratory and stereotyped behavior as well as spatial memory and learning. TCB-2 and 25CN-NBOH increased the BDNF mRNA level. All 5-HT2A agonists increased the proBDNF level but failed to alter the mature BDNF protein level. TrkB and p75NTR mRNA levels were affected by all utilized agonists. All drugs decreased the total level as well as membrane TrkB protein one indicating downregulation of TrkB receptors. All agonists decreased the membrane p75NTR protein level. Thus, we have shown for the first time that the chronic activation of the 5-HT2A receptor with agonists has affected the BDNF system almost on all levels-transcription, proBDNF production, TrkB and p75NTR receptors' level. The obtained data suggested possible suppression in BDNF-TrkB signaling under chronic treatment with 5-HT2A agonists.

On the interaction between BDNF and serotonin systems: The effects of long-term ethanol consumption in mice.

We investigated the effect of chronic (6 weeks) consumption of 10% alcohol on the principal elements of BDNF (BDNF, proBDNF, p75, and TrkB receptors) and 5-HT (5-HT, 5-HIAA, tryptophan hydroxylase-2 [Tph-2], 5-HT transporter [5-HTT], 5-HT1A, 5-HT2A, and 5-HT7 receptors) systems in the brain of C57Bl/6 mice. BDNF mRNA level in the raphe nuclei area and BDNF protein level in the hippocampus were lowered in ethanol-treated mice. The increase in proBDNF protein level in the raphe nuclei area, cortex, and amygdala and the increase of p75 receptor protein levels in the raphe nuclei area were revealed after ethanol exposure. Alcohol intake reduced the protein level and increased the activity of Tph-2, the key enzyme for serotonin biosynthesis in the brain, and increased the main 5-HT metabolite 5-HIAA level and 5-HIAA/5-НТ ratio as well as the 5-HT7 receptor mRNA level in the raphe nuclei area. In the cortex, 5-HT2A receptor protein level was reduced, and 5-HIAA/5-HT ratio was increased. These data showed considerable impact of alcoholization on the BDNF system, resulting in proBDNF and p75 receptor expression enhancement. Alcohol-induced changes in BDNF and 5-HT systems were revealed in the raphe nuclei area where the majority of the cell bodies of the 5-HT neurons are localized, as well as in the cortex, hippocampus, and amygdala. Our data suggest that the BDNF/5-HT interaction contributes to the mechanism underlying chronic alcohol-induced neurodegenerative disorders.

Genetically defined fear-induced aggression: Focus on BDNF and its receptors.

Brain-derived neurotrophic factor (BDNF), its precursor proBDNF, BDNF pro-peptide, BDNF mRNA levels, as well as TrkB and p75NTR receptors mRNA and protein levels, were studied in the brain of rats, selectively bred for more than 85 generations for either the high level or the lack of fear-induced aggressive behavior. Furthermore, we have found that rats of aggressive strain demonstrated both high level of aggression toward humans and increased amplitude of acoustic startle response compared to rats selectively bred for the lack of fear-induced aggression. Significant increase in the BDNF mRNA, mature BDNF and proBDNF protein levels in the raphe nuclei (RN), hippocampus (Hc), nucleus accumbens (NAcc), amygdala, striatum and hypothalamus (Ht) of aggressive rats was revealed. The BDNF/proBDNF ratio was significantly reduced in the Hc and NAcc of highly aggressive rats suggesting prevalence of the proBDNF in these structures. In the Hc and frontal cortex (FC) of aggressive rats, the level of the full-length TrkB (TrkB-FL) receptor form was decreased, whereas the truncated TrkB (TrkB-T) protein level was increased in the RN, FC, substantia nigra and Ht. The TrkB-FL/TrkB-T ratio was significantly decreased in highly aggressive rats suggesting TrkB-T is predominant in highly aggressive rats. The p75NTR expression was slightly changed in majority of studied brain structures of aggressive rats. The data indicate the BDNF system in the brain of aggressive and nonaggressive animals is extremely different at all levels, from transcription to reception, suggesting significant role of BDNF system in the development of highly aggressive phenotype.

Role of glial cell line-derived neurotrophic factor in the pathogenesis and treatment of mood disorders.

Glial cell line-derived neurotrophic factor (GDNF) is widely recognized as a survival factor for dopaminergic neurons, but GDNF has also been shown to promote development, differentiation, and protection of other central nervous system neurons and was thought to play an important role in various neuropsychiatric disorders. Severe mood disorders, such as primarily major depressive disorder and bipolar affective disorder, attract particular attention. These psychopathologies are characterized by structural alterations accompanied by the dysregulation of neuroprotective and neurotrophic signaling mechanisms required for the maturation, growth, and survival of neurons and glia. The main objective of this review is to summarize the recent findings and evaluate the potential role of GDNF in the pathogenesis and treatment of mood disorders. Specifically, it describes (1) the implication of GDNF in the mechanism of depression and in the effect of antidepressant drugs and mood stabilizers and (2) the interrelation between GDNF and brain neurotransmitters, playing a key role in the pathogenesis of depression. This review provides converging lines of evidence that (1) brain GDNF contributes to the mechanism underlying depressive disorders and the effect of antidepressants and mood stabilizers and (2) there is a cross-talk between GDNF and neurotransmitters representing a feedback system: GDNF-neurotransmitters and neurotransmitters-GDNF.

Glial cell line-derived neurotrophic factor in genetically defined fear-induced aggression.

Glial cell line-derived neurotrophic factor (GDNF) plays an important role in maintenance of neuronal system throughout life. However, there is a lack of data on the involvement of GDNF in the regulation of different kinds of behavior. In this study, GDNF, its precursor (proGDNF) and GDNF mRNA levels were investigated in the brain of rats selectively bred for 85 generations for either high level or for the lack of affective aggressiveness toward human. It was found that GDNF mRNA level was decreased in the frontal cortex, increased in the raphe nuclei area of the midbrain of aggressive rats compared to tame animals and was not detected in the amygdala and hypothalamus. The level of proGDNF was reduced in the raphe nuclei area of the midbrain of highly aggressive rats and was not detected in the striatum, nucleus accumbens of investigated animals. Two forms of mature GDNF - monomer and dimer - were revealed. GDNF monomer level was increased in the raphe nuclei area, substantia nigra and amygdala of aggressive rats and it was not found in the frontal cortex and nucleus accumbens of investigated rats. Dimer GDNF level was found in all investigated brain structures. It was reduced in the hippocampus and increased in amygdala of highly aggressive rats. Thus, considerable structure-specific differences in GDNF expression between highly aggressive and nonaggressive rats were shown. The data suggested the implication of both mature GDNF monomer and dimer as well as proGDNF in the mechanism underlying genetically defined aggressiveness.

5-HT1A receptor gene silencers Freud-1 and Freud-2 are differently expressed in the brain of rats with genetically determined high level of fear-induced aggression or its absence.

Serotonin 5-HT1A receptor is known to play a crucial role in the mechanisms of genetically defined aggression. In its turn, 5-HT1A receptor functional state is under control of multiple factors. Among others, transcriptional factors Freud-1 and Freud-2 are known to be involved in the repression of 5-HT1A receptor gene expression. However, implication of these factors in the regulation of behavior is unclear. Here, we investigated the expression of 5-HT1A receptor and silencers Freud-1 and Freud-2 in the brain of rats selectively bred for 85 generations for either high level of fear-induced aggression or its absence. It was shown that Freud-1 and Freud-2 levels were different in aggressive and nonaggressive animals. Freud-1 protein level was decreased in the hippocampus, whereas Freud-2 protein level was increased in the frontal cortex of highly aggressive rats. There no differences in 5-HT1A receptor gene expression were found in the brains of highly aggressive and nonaggressive rats. However, 5-HT1A receptor protein level was decreased in the midbrain and increased in the hippocampus of highly aggressive rats. These data showed the involvement of Freud-1 and Freud-2 in the regulation of genetically defined fear-induced aggression. However, these silencers do not affect transcription of the 5-HT1A receptor gene in the investigated rats. Our data indicate the implication of posttranscriptional rather than transcriptional regulation of 5-HT1A receptor functional state in the mechanisms of genetically determined aggressive behavior. On the other hand, the implication of other transcriptional regulators for 5-HT1A receptor gene in the mechanisms of genetically defined aggression could be suggested.

Brain-derived neurotrophic factor (BDNF) and its precursor (proBDNF) in genetically defined fear-induced aggression.

The brain-derived neurotrophic factor (BDNF), its precursor (proBDNF) and BDNF mRNA levels were studied in the brain of wild rats selectively bred for more than 70 generations for either high level or for the lack of affective aggressiveness towards man. Significant increase of BDNF mRNA level in the frontal cortex and increase of BDNF level in the hippocampus of aggressive rats was revealed. In the midbrain and hippocampus of aggressive rats proBDNF level was increased, whereas BDNF/proBDNF ratio was reduced suggesting the prevalence and increased influence of proBDNF in highly aggressive rats. In the frontal cortex, proBDNF level in aggressive rats was decreased. Thus, considerable structure-specific differences in BDNF and proBDNF levels as well as in BDNF gene expression between highly aggressive and nonaggressive rats were shown. The data suggested the implication of BDNF and its precursor proBDNF in the mechanism of aggressiveness and in the creation of either aggressive or nonaggressive phenotype.