Early-life stress affects peripheral, blood-brain barrier, and brain responses to immune challenge in juvenile and adult rats.

Early-life stress (ELS) may affect brain maturation and neuroimmune interactions and, consequently, the inflammatory response to subsequent environmental factors later in life. Recently, the coexistence of blood-brain barrier (BBB) dysfunction and inflammation has been implicated in the etiology and progression of mental and/or neurodegenerative diseases. There are sex differences in the prevalence and outcomes of these disorders. The number of studies reporting the effects of ELS and sex on BBB functioning and neuroinflammatory processes in response to immune challenge is very limited, and the data are inconsistent. In the present study, we examined whether ELS, based on the maternal separation (MS) paradigm in rats, can condition male and female subjects to subsequent lipopolysaccharide (LPS)-induced immune challenge in juvenility or adulthood. Twenty-four hours after acute LPS injection, serum proinflammatory cytokines were measured, and BBB permeability in the medial prefrontal cortex (mPFC) and hippocampus (HP) was evaluated. Additionally, the mRNA expression of neuroinflammatory markers and BBB-related genes was also studied. We found that a single LPS challenge induced a proinflammatory response both in the periphery and in the mPFC and HP and increased BBB permeability in a sex-dependent fashion. Moreover, MS enhanced the neuroinflammatory response to LPS challenge in males (especially juveniles), whereas MS females showed no difference or a blunted central response to LPS compared with control females, mainly during adulthood. These results suggest that ELS may precondition individuals to subsequent environmental factors later in life in a sex-specific manner and potentially determine their susceptibility or resilience to mental and/or neurodegenerative diseases.

Early-life stress affects the structural and functional plasticity of the medial prefrontal cortex in adolescent rats.

Early life experiences are crucial factors that shape brain development and function due to their ability to induce structural and functional plasticity. Among these experiences, early-life stress (ELS) is known to interfere with brain development and maturation, increasing the risk of future psychopathologies, including depression, anxiety, and personality disorders. Moreover, ELS may contribute to the emergence of these psychopathologies during adolescence. In this present study, we investigated the effects of ELS, in the form of maternal separation (MS), on the structural and functional plasticity of the medial prefrontal cortex (mPFC) and anxiety-like behavior in adolescent male rats. We found that the MS procedure resulted in disturbances in mother-pup interactions that lasted until weaning and were most strongly demonstrated by increases in nursing behavior. Moreover, MS caused atrophy of the basal dendritic tree and reduced spine density on both the apical and basal dendrites in layer II/III pyramidal neurons of the mPFC. The structural changes were accompanied by an impairment of long-term potentiation processes and increased expression of key proteins, specifically glutamate receptor 1, glutamate receptor 2, postsynaptic density protein 95, αCa(2+) /calmodulin-dependent protein kinase II and αCa(2+)/calmodulin-dependent protein kinase II phosphorylated at residue Thr305, that are engaged in long-term potentiation induction and maintenance in the mPFC. We also found that the MS animals were more anxious in the light/dark exploration test. The results of this study indicate that ELS has a significant impact on the structural and functional plasticity of the mPFC in adolescents. ELS-induced adaptive plasticity may underlie the pathomechanisms of some early-onset psychopathologies observed in adolescents.

Modulation of the endoplasmic reticulum stress and unfolded protein response mitigates the behavioral effects of early-life stress.

BACKGROUND: Early-life stress (ELS) affects brain development and increases the risk of mental disorders associated with the dysfunction of the medial prefrontal cortex (mPFC). The mechanisms of ELS action are not well understood. Endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) are cellular processes involved in brain maturation through the regulation of pro-survival or proapoptotic processes. We hypothesized that ER stress and the UPR in the mPFC are involved in the neurobiology of ELS. METHODS: We performed a maternal separation (MS) procedure from postnatal days 1 to 14 in rats. Before each MS, pups were injected with an inhibitor of ER stress, salubrinal or a vehicle. The mRNA and protein expression of UPR and apoptotic markers were evaluated in the mPFC using RT-qPCR and Western blot methods, respectively. We also estimated the numbers of neurons and glial cells using stereological methods. Additionally, we assessed behavioral phenotypes related to fear, anhedonia and response to psychostimulants. RESULTS: MS slightly enhanced the activation of the UPR in juveniles and modulated the expression of apoptotic markers in juveniles and preadolescents but not in adults. Additionally, MS did not affect the numbers of neurons and glial cells at any age. Both salubrinal and vehicle blunted the expression of UPR markers in juvenile and preadolescent MS rats, often in a treatment-specific manner. Moreover, salubrinal and vehicle generally alleviated the behavioral effects of MS in preadolescent and adult rats. CONCLUSIONS: Modulation of ER stress and UPR processes may potentially underlie susceptibility or resilience to ELS.

Effects of chronic fluoxetine treatment on anxiety- and depressive-like behaviors in adolescent rodents - systematic review and meta-analysis.

BACKGROUND: Drugs prescribed for psychiatric disorders in adolescence should be studied very extensively since they can affect developing and thus highly plastic brain differently than they affect the adult brain. Therefore, we aimed to summarize animal studies reporting the behavioral consequences of chronic exposure to the most widely prescribed antidepressant drug among adolescents i.e., fluoxetine. METHODS: Electronic databases (Medline via Pubmed, Web of Science Core Collection, ScienceDirect) were systematically searched until April 12, 2022, for published, peer-reviewed, controlled trials concerning the effects of chronic fluoxetine administration vs. vehicle on anxiety and depression measures in naïve and stress-exposed adolescent rodents. All of the relevant studies were selected and critically appraised, and a meta-analysis of eligible studies was performed. RESULTS: A total of 18 studies were included in the meta-analysis. In naïve animals, chronic adolescent fluoxetine administration showed dose-related anxiogenic-like effects, measured as a reduction in time spent in the open arms of the elevated plus maze. No significant effects of chronic adolescent fluoxetine on depression-like behavior were reported in naïve animals, while in stress-exposed rodents chronic adolescent fluoxetine significantly decreased immobility time in the forced swim test compared to vehicle. CONCLUSIONS: These results suggest that although chronic fluoxetine treatment proves positive effects in animal models of depression, it may simultaneously increase anxiety in adolescent animals in a dose-related manner. Although the clinical implications of the data should be interpreted with extreme caution, adolescent patients under fluoxetine treatment should be closely monitored.

Effects of early-life stress and sex on blood-brain barrier permeability and integrity in juvenile and adult rats.

Early-life stress (ELS) is considered a relevant etiological factor for neurodegenerative and mental disorders. In the present study, we hypothesized that ELS may persistently and sex dependently influence blood-brain barrier (BBB) integrity and function during critical periods of brain development and consequently determine susceptibility to and sex-related prevalence of chronic diseases in adult life. We used the maternal separation (MS) procedure in rats to model ELS and evaluated BBB permeability and gene expression of selected tight junction (TJ) proteins, glucose transporter type 1 (Slc2a1) and aquaporin 4 (Aqp4) in the medial prefrontal cortex (mPFC), dorsal striatum (dSTR) and hippocampus of juvenile and adult rats. Serum concentrations of a peripheral marker of BBB function (S100ß) and proinflammatory cytokines were also assessed. We observed developmental sealing of the BBB and sex differences in the permeability of the BBB and the mRNA expression of TJ proteins and Slc2a1. Adult females showed lower BBB permeability and higher levels of Cldn3, Cldn5, Ocln, and Slc2a1 in the mPFC and dSTR than males. MS temporarily increased BBB permeability in the dSTR of juvenile males and affected mRNA expression of the majority of studied proteins related to BBB function in age-, region- and sex-dependent manners. Additionally, MS sex dependently decreased serum S100ß levels and did not affect proinflammatory cytokine concentrations. In general, our study did not reveal a clear or strong negative effect of MS on BBB integrity. However, the results suggest that ELS may induce adaptive/maladaptive changes or compensatory mechanisms within the BBB of unknown yet consequences.

A Search for Biomarkers of Early-life Stress-related Psychopathology: Focus on 70-kDa Heat Shock Proteins.

Clinical studies clearly indicate that early-life stress (ELS) may cause physical and mental health problems later in life. Therefore, the identification of universal biomarkers of ELS-related diseases is very important. The 70-kDa heat shock proteins (HSP70s), specifically HSPA5 and HSPA1B, have been recently shown to be potentially associated with occurrence of anxiety, mood disorders, and schizophrenia; thus, we hypothesized that HSP70s are potential candidate biomarkers of ELS-induced psychopathologies. A maternal separation (MS) procedure in rats was used to model ELS, and the expression of HSPA5 and HSPA1B was investigated in the blood, medial prefrontal cortex (mPFC), and hippocampus of juvenile, preadolescent, and adult animals. We also studied the effects of MS on the long-term potentiation (LTP) and behavioral phenotypes of adult rats. We found that MS enhanced the expression of HSPA1B mRNA in the blood and mPFC of juvenile and preadolescent rats. This increase was accompanied by an increase in the HSPA1A/1B protein levels in the mPFC and hippocampus of juvenile rats that persisted in the mPFC until adulthood. MS juvenile and adult rats showed enhanced HSPA5 mRNA expression in the blood and increased HSPA5 protein expression in the mPFC (juveniles) and hippocampus (adults). Concurrently, MS adult rats exhibited aberrations in LTP in the mPFC and hippocampus and a less anxious behavioral phenotype. These results indicate that MS may produce enduring overexpression of HSPA1B and HSPA5 in the brain and blood. Therefore, both HSP70 family members may be potential candidate peripheral and brain biomarkers of ELS-induced changes in brain functioning.

1MeTIQ and olanzapine, despite their neurochemical impact, did not ameliorate performance in fear conditioning and social interaction tests in an MK-801 rat model of schizophrenia.

BACKGROUND: The aim of the present study was to evaluate the effect of 1MeTIQ on fear memory and social interaction in an MK-801-induced model of schizophrenia. The results obtained after administration of 1MeTIQ were compared with those obtained with olanzapine, an antipsychotic drug. METHODS: Sprague-Dawley rats received a single injection of MK-801 to induce behavioral disorders. 1MeTIQ was given either acutely in a single dose or chronically for 7 consecutive days. Olanzapine was administered once. In groups receiving combined treatments, 1MeTIQ or olanzapine was administered 20 min before MK-801 injection. Contextual fear conditioning was used to assess disturbances in fear memory (FM), and the sociability of the rats was measured in the social interaction test (SIT). Biochemical analysis was carried out to evaluate monoamine levels in selected brain structures after treatment. RESULTS: Our results are focused mainly on data obtained from neurochemical studies, demonstrating that 1MeTIQ inhibited the MK-801-induced reduction in dopamine levels in the frontal cortex and increased the 5-HT concentration. The behavioral tests revealed that acute administration of MK-801 caused disturbances in both the FM and SIT tests, while neither 1MeTIQ nor olanzapine reversed these deficits. CONCLUSION: 1MeTIQ, although pharmacologically effective (i.e., it reverses MK-801-induced changes in monoamine activity), did not influence MK-801-induced social and cognitive deficits. Thus, our FM tests and SIT did not support the main pharmacological hypotheses that focus on dopamine system stabilization and dopamine-serotonin system interactions as probable mechanisms for inhibiting the negative symptoms of schizophrenia.

Simultaneous activation of mGlu2 and muscarinic receptors reverses MK-801-induced cognitive decline in rodents.

The activity of an allosteric agonist of muscarinic M1 receptor, VU0357017, and a positive allosteric modulator (PAM) of M5 receptor, VU0238429, were investigated alone or in combination with the mGlu2 receptor PAM, LY487379 using the following behavioural tests: prepulse inhibition (PPI), novel object recognition (NOR), and spatial delayed alternation (SDA). VU0357017 (10 and 20 mg/kg) and VU0238429 (5 and 10 mg/kg) reversed deficits in PPI while VU0238429 (2.5 and 5 mg/kg) was effective in SDA. The simultaneous administration of subeffective doses of M1 or M5 activators (5, 1, or 0.25 mg/kg) with LY487379 (0.5 mg/kg) induced the same effect as that observed for the active dose of each compound. Selective M1 or M5 receptor blockers antagonized the effect exerted by these combinations, and pharmacokinetic studies confirmed independent transport through the blood-brain barrier. The expression of both receptors (M1 and M5) was established in brain structures involved in cognition (neocortex, hippocampus, and entorhinal cortex) in both the rat and the mouse brains by immunofluorescence staining. Specifically, double neuronal staining of mGlu2-M1 and mGlu2-M5 receptors was observed in many areas of the rat brain, while the number of double-stained mGlu2-M1 receptors was moderate in the mouse brain with no mGlu2-M5 colocalization. Finally, the combined administration of subeffective doses of the compounds did not alter prolactin levels or motor coordination, in contrast to the compounds given alone at the highest dose or in combination with standard neuroleptics.

The Trace Kynurenine, Cinnabarinic Acid, Displays Potent Antipsychotic-Like Activity in Mice and Its Levels Are Reduced in the Prefrontal Cortex of Individuals Affected by Schizophrenia.

Cinnabarinic acid (CA) is a kynurenine metabolite that activates mGlu4 metabotropic glutamate receptors. Using a highly sensitive ultra-performance liquid chromatography/tandem mass spectrometry (UPLC/MS-MS) method, we found that CA is present in trace amounts in human brain tissue. CA levels were largely reduced in the prefrontal cortex (PFC) of individuals affected by schizophrenia. This reduction did not correlate with age, sex, duration of the disease, and duration and type of antipsychotic medication and might, therefore, represent a trait of schizophrenia. Interestingly, systemic treatment with low doses of CA (<1 mg/kg, i.p.) showed robust efficacy in several behavioral tests useful to study antipsychotic-like activity in mice and rats and attenuated MK-801-evoked glutamate release. CA failed to display antipsychotic-like activity and inhibit excitatory synaptic transmission in mice lacking mGlu4 receptors. These findings suggest that CA is a potent endogenous antipsychotic-like molecule and reduced CA levels in the PFC might contribute to the pathophysiology of schizophrenia.

The impact of early-life stress on corticosteroid carrier protein levels and 11ß-hydroxysteroid dehydrogenase 1 expression in adolescent rats.

BACKGROUND: Corticosteroid-binding globulin (CBG), albumin and 11ß-hydroxysteroid dehydrogenase (11ß-HSD) enzymes play crucial roles in the bioavailability of glucocorticoids. Downstream of the adrenal glands, these proteins affect glucocorticoid levels in target tissues. Early-life stress (ELS) is known to program glucocorticoid action at many levels. The effects of ELS on the concentrations and synthesis of CBG and albumin and on the expression of 11ß-HSD remain unclear. METHODS: The maternal separation (MS) procedure in rats on postnatal days 1-14 was used as a model of ELS. On postnatal day 35 (adolescence), the serum corticosterone, CBG and albumin concentrations of male rats were measured by ELISA, while the mRNA and protein levels of CBG, albumin and 11ß-HSD1 in the liver and brain were examined by RT-qPCR and Western blot, respectively. RESULTS: Under basal conditions, MS rats displayed lower levels of serum CBG and albumin. However, MS did not affect CBG or albumin synthesis in the liver, suggesting that the half-life and/or secretion of these proteins were influenced by MS. Additionally, MS rats showed increased protein expression of 11ß-HSD1, specifically in the medial prefrontal cortex. CONCLUSIONS: These results indicate that ELS may potentially program glucocorticoid action through its effects on glucocorticoid bioavailability in tissues.

Maternal separation disturbs postnatal development of the medial prefrontal cortex and affects the number of neurons and glial cells in adolescent rats.

Adolescence is a period of extensive brain maturation. In particular, the regions of the medial prefrontal cortex (mPFC) undergo intense structural and functional refinement during adolescence. Disturbances in mPFC maturation have been implicated in the emergence of multiple psychopathologies during adolescence. One of the essential risk factors for the development of mental illness in adolescence is early-life stress (ELS), which may interfere with brain maturation. However, knowledge of the mechanisms by which ELS affects mPFC maturation and functioning in adolescents is very limited. In the present study, we applied a maternal separation (MS) procedure in rats to model ELS and studied its effect on the number of neurons and glial cells in the prelimbic region of the mPFC (PLC) of adolescent rats. Moreover, the expression of markers of cell proliferation and apoptosis was also studied. We found that MS rats had more neurons, astrocytes, and NG2-glial cells in the PLC. In contrast, the number of microglial cells was reduced in MS rats. These changes were accompanied by the decreased expression of proapoptotic genes and the increased expression of some prosurvival genes. Concurrently, MS did not affect cell proliferation in adolescents. Moreover, MS induced anxiety-like behaviors, but not anhedonic-like behavior, in adolescents. These results suggest that ELS may disturb neurodevelopmental apoptosis of neurons and early-postnatal proliferation and/or apoptosis of different populations of glial cells in the PLC. ELS-induced aberrations in the postnatal maturation of the PLC may affect cortical network organization and functioning and determine vulnerability to psychopathologies in adolescents.

Cocaine decreases the expression of PSA-NCAM protein and attenuates long-term potentiation via glucocorticoid receptors in the rat dentate gyrus.

The present study investigated a potential role for glucocorticoid (GR) and mineralocorticoid (MR) receptors in the detrimental effects of single cocaine (COC) administration on both the number of polysialylated neural cell adhesion molecule (PSA-NCAM)-positive neurons and the induction of long-term potentiation (LTP) in the rat dentate gyrus (DG). The effects of COC (15 mg/kg i.p.) on the number of PSA-NCAM-positive neurons and the induction of LTP observed 2 days after COC administration were abolished either by depleting circulating corticosterone after administration of metyrapone (100 mg/kg s.c. given 3 h before COC) or by pharmacologically blocking GRs using mifepristone (RU 38486, 10 mg/kg s.c. given 1 h before COC). Administration of the MR blocker spironolactone (50 mg/kg s.c. given 1 h before COC) did not alter the effects of COC on the number of PSA-NCAM-positive neurons or LTP induction. Results have also shown that COC does not change the rate of cell proliferation, as measured by the presence of Ki-67 and the incorporation of bromodeoxyuridine (100 mg/kg i.p. given 2 h after COC) into the newly born cells in the DG 2 days after COC administration. Finally, we observed that GRs colocalized with some, but not all, PSA-NCAM-positive neurons, whereas MRs showed no colocalization with neurons positive for PSA-NCAM in the DG. These data indicate that a single dose of COC may arrest hippocampal susceptibility to plastic changes and lead to functional impairments through the alteration of hippocampal structure and the formation of memory traces.

Detrimental effect of postnatal blockade of N-methyl-D-aspartate receptors on sensorimotor gating is reversed by neuroleptic drugs.

Postnatal hypofunction of N-methyl-D-aspartate (NMDA) receptors leads to several behavioral deficits in adult rats resembling deficits typical of schizophrenia-like deficits of sensorimotor gating. Thus far, it is not known whether the above disruptions are sensitive to neuroleptic drugs. In order to verify the above model in pharmacological terms, we investigated whether deficits in the sensorimotor gating evoked by administration of NMDA receptor antagonists in the postnatal period is sensitive to neuroleptic drugs. We also investigated whether such treatment evoked alterations in the expression of dopamine D(1), D(2) and D(3) receptors in the nucleus accumbens, a key structure for dopamine-dependent alterations in sensorimotor gating. CGP 40116, a competitive antagonist of NMDA receptors was given in doses of 1.25 mg/kg on days 1, 3, 6 and 9; 2.5 mg/kg on days 12, 15 and 18; and 5 mg/kg on day 21 (all injections were sc). The efficacy of sensorimotor gating was tested on rats at the age of 60 days using a prepulse-induced inhibition of the startle reflex. In order to measure the expression of dopamine D(1), D(2) and D(3) receptors, we used quantitative autoradiography and tritiated ligands i.e. [(3)H]-SCH 23390, [(3)H]-Spiperone and [(3)H]-7-OH-DPAT, respectively. Haloperidol (0.1 mg/kg, sc), risperidone (1.0 mg/kg, sc) and clozapine (2.5 mg/kg, sc) reversed deficits of sensorimotor gating observed in adult rats evoked by the postnatal administration of CGP 40116. We also observed enhanced density of dopamine D(3), but not D(1) and D(2) receptors in the nucleus accumbens of CGP40116 treated rats. It is concluded that models of cognitive dysfunction, typical for schizophrenia based on postnatal administration of NMDA receptor antagonists, are sensitive to neuroleptic drugs and possibly not dependent on alteration in the density of dopaminergic receptors.

Previous Early-life Stress Modifies Acute Corticosterone-induced Synaptic Plasticity in the Medial Prefrontal Cortex of Adolescent Rats.

Stress can either strengthen coping strategies or enhance the risk of depression and anxiety. Synaptic plasticity is one of the key brain functions that can be affected by stress. We have previously shown that early-life stress in the form of maternal separation (MS) impairs functional synaptic plasticity in the medial prefrontal cortex (mPFC), i.e., long-term potentiation (LTP), in adolescent rats. It has been postulated that a previous experience of prolonged stress can modify the response to a subsequent acute stress challenge and influence coping strategies. Therefore, in the present study, we examined how previous MS experience influenced acute stress-induced changes in the LTP and expression of genes and proteins engaged in synaptic plasticity in the mPFC of adolescent rats. To mimic acute stress, we applied acute injections of corticosterone (CORT) and its vehicle (VEH). In control rats, acute CORT injection enhanced LTP in the mPFC. In contrast, MS rats generally exhibited an impairment of LTP that was not further affected by CORT. Moreover, for many studied parameters, such as induction of cFos and Arc mRNA and protein and activation of BDNF, GDNF and NCAM mRNA, MS rats showed diminished, vague or absent responses to acute VEH/CORT compared with those of control rats. These results suggest that previous early-life stress experiences may induce adaptive plasticity within the mPFC, which influences the response to acute stress challenge and coping strategies in adolescents. Depending on the specific environmental context, this phenomenon may lead to either future vulnerability or future resilience to stress-related psychopathologies.

Negative Allosteric Modulators of mGlu7 Receptor as Putative Antipsychotic Drugs.

The data concerning antipsychotic-like activity of negative allosteric modulators (NAMs)/antagonists of mGlu7 receptors are limited. The only available ligands for this receptor are MMPIP and ADX71743. In the present studies, we used stable cell line expressing mGlu7 receptor and it was shown that both compounds dose-dependently potentiated forskolin elevated cAMP concentration in the T-REx 293 cells, showing their inverse agonist properties. Subsequently, pharmacokinetic studies were performed. Both compounds were given intraperitoneally (i.p.) at the dose of 10 mg/kg and reached Cmax 0.25-0.5 h after administration, and then they declined rapidly, ADX71743 being almost undetectable 2 h after administration, while the concentration of MMPIP was still observed, suggesting that the concentration of MMPIP was more stable. Finally, we investigated the role of both mGlu7 receptor NAMs in animal models of schizophrenia. Behavioral tests commonly used in antipsychotic drug discovery were conducted. Both tested compounds dose-dependently inhibited MK-801-induced hyperactivity (MMPIP at 15 mg/kg; ADX at 5 and 15 mg/kg) and DOI-induced head twitches (MMPIP at 5, 10, 15 mg/kg; ADX at 2.5, 5, 10 mg/kg). Moreover, the same effects were noticed in novel object recognition test, where MMPIP (5, 10, 15 mg/kg) and ADX71743 (1, 5, 15 mg/kg) reversed MK-801-induced disturbances. In the social interaction test, antipsychotic activity was observed only for ADX71743 (5, 15 mg/kg). ADX71743 at the dose 2.5 mg/kg reversed MK-801-induced disruption in prepulse inhibition while MMPIP at 10 mg/kg reversed MK-801-induced disruption in spatial delayed alternation. The present studies showed that mGlu7 receptor may be considered as a putative target for antipsychotic drugs, though more studies are needed due to limited number of available ligands.

Acute activation of CB1 cannabinoid receptors transiently decreases PSA-NCAM expression in the dentate gyrus of the rat hippocampus.

Recent evidence indicates that the polysialylated neural cell adhesion molecule (PSA-NCAM) is involved in hippocampal plasticity. On the other hand, CB1 receptor activation is known to disturb some hippocampal processes involving plastic changes, such as learning and memory. Therefore, the present study investigated the effect of HU-210, a CB1 receptor agonist, on the expression of PSA-NCAM protein in the dentate gyrus (DG) and CA3 region of the rat hippocampus. It was found that at a dose of 0.1 mg/kg i.p. of HU-210, the number of PSA-NCAM immunoreactive (IR) cells in the DG declined in a time-dependent manner. The decrease in PSA-NCAM expression was observed at 1 and 2 days (ca. 21% and 30%, respectively), but not after 4 h and 4 days following HU-210 administration. However, HU-210 treatment did not change the length density of PSA-NCAM immunopositive processes in CA3 mossy fibers at all the time points measured. The effect observed in the DG on day 2 was blocked by AM-251 (1 mg/kg, i.p.), a CB1 receptor antagonist, given 30 min before HU-210. Neither the number of Ki-67 (IR) cells (a marker of proliferation) nor the number of doublecortin-IR cells (a marker of immature neurons) was affected by HU-210 (0.1 mg/kg, i.p.) treatment at any of the time points. An analysis of co-localization of CB1 receptor protein with PSA-NCAM protein revealed that both proteins were not present in the same population of neurons in the subgranular layer of the DG. The observed changes in PSA-NCAM expression were not related to the reduction of proliferation or differentiation of newly born cells, but were possible due to alternations in the synaptic activity in the DG. However, such alteration in the PSA-NCAM expression may change the timing of the functional maturation of newly born neurons. Moreover, the above finding suggests that acute activation of CB1 receptors may result in the stiffening of the hippocampal structure and susceptibility to plastic changes and may lead to functional impairment governed by alterations in the hippocampal structure.

The effects of early-life stress on dopamine system function in adolescent female rats.

During adolescence, many neural systems, including the dopamine system, undergo essential remodeling and maturation. It is well known that early-life stress (ELS) increases the risk for many psychopathologies during adolescence and adulthood. It is hypothesized that ELS interferes with the maturation of the dopamine system. There is a sex bias in the prevalence of stress-related mental disorders. Information regarding the effects of ELS on brain functioning in females is very limited. In the current study, maternal separation (MS) procedures were carried out to study the effects of ELS on dopamine system functioning in adolescent female rats. Our study showed that MS increased the density of tyrosine hydroxylase immunoreactive fibers in the prelimbic cortex (PLC) and nucleus accumbens (Acb). These changes were accompanied by a decrease in the level of D5 receptor mRNA and an increase in D2 receptor mRNA expression in the PLC of MS females. Conversely, D1 and D5 receptor mRNA levels were augmented in the caudate putamen (CPu), while the expression of the D3 dopamine receptor transcript was reduced in MS females. Additionally, in the Acb, MS elicited a decrease in D2 receptor mRNA expression. At the behavioral level, MS increased apomorphine-induced locomotion; however, it did not change locomotor responses to selective D1/D5 receptor agonist and attenuated D2/D3 receptor agonist-triggered locomotion. Moreover, MS decreased D1/D5 receptor agonist-induced grooming behavior. These results indicate that ELS disrupts dopamine receptor function in the PLC and basal ganglia during adolescence in females and may predispose them to psychopathologies during adolescence and adulthood.

Regulation of alpha-synuclein expression in limbic and motor brain regions of morphine-treated mice.

Chronic exposure to opiates produces dependence and addiction, which may result from neuroadaptations in the dopaminergic reward pathway and its target brain regions. The neuronal protein alpha-synuclein has been implicated in neuronal plasticity and proposed to serve as a negative regulator of dopamine neurotransmission. Thus, alpha-synuclein could mediate some effects of opiates in the brain. The present study investigated the influence of acute and chronic morphine administration on alpha-synuclein mRNA and protein expression in the brains of mice. Downregulation of alpha-synuclein mRNA was observed in the basolateral amygdala, dorsal striatum, nucleus accumbens, and ventral tegmental area of mice withdrawn from chronic morphine treatment. The changes were the most pronounced after longer periods of withdrawal (48 h). In contrast, levels of alpha-synuclein protein, as assessed by Western blotting, were significantly increased in the amygdala and striatum/accumbens (but not in the mesencephalon) of morphine-withdrawn mice. In both brain regions, levels of alpha-synuclein were elevated for as long as 2 weeks after treatment cessation. Because alpha-synuclein is a presynaptic protein, the detected opposite changes in its mRNA and protein levels are likely to take place in different populations of projection neurons whose somata are in different brain areas. Axonal localization of alpha-synuclein was confirmed by immunofluorescent labeling. An attempt to identify postsynaptic neurons innervated by alpha-synuclein-containing axon terminals revealed their selective apposition to calbindin D28K-negative projection neurons in the basolateral amygdala. The observed changes in alpha-synuclein levels are discussed in connection with their putative role in mediating suppression of dopaminergic neurotransmission during opiate withdrawal.

Acute and repeated cocaine induces alterations in FosB/DeltaFosB expression in the paraventricular nucleus of the hypothalamus.

Apart from activation of the brain reward system, cocaine administration influences the activity of the hypothalamo-pituitary-adrenal (HPA) axis by affecting CRH neurons in the paraventricular nucleus of the hypothalamus (PVN). In order to find a molecular mechanism of cocaine-evoked effects in the PVN, in the present study, we investigated the impact of cocaine on the expression of FosB/DeltaFosB transcription factors in the PVN. Using an immunohistochemical method, we found that acute cocaine treatment (25 mg/kg) induced a relatively long-lasting (at least 72 h) expression of FosB/DeltaFosB in the PVN, whereas repeated cocaine administration (25 mg/kg, once daily for 5 consecutive days) caused accumulation of FosB/DeltaFosB in the PVN. The latter observation was further confirmed by the Western blot technique which revealed that repeated exposure to cocaine specifically increased the expression of a stable isoform of DeltaFosB (35 kDa). Using a double-labeling immunofluorescent method, it was established that FosB/DeltaFosB proteins induced by repeated cocaine treatment were present in a small population of CRF-immunoreactive neurons of the PVN. Furthermore, it was found that pretreatment with the specific antagonist of dopamine D1-like receptors SCH 23390 (1 mg/kg) attenuated the expression and accumulation of FosB/DeltaFosB in the PVN, evoked by repeated cocaine administration. Although functional consequences of the above effects for the process of addiction remain to be established, the obtained results indicate that cocaine administration can produce relatively long-lasting changes in the expression of FosB/DeltaFosB transcription factors in PVN neurons (in some populations of CRF-immunoreactive neurons, among others) and that dopamine D1-like receptors are involved in the above effects. Finally, it is proposed that the long-lasting expression as well as the accumulation of DeltaFosB in the PVN may constitute a molecular basis underlying adaptive changes occurring in the HPA axis after relatively high doses of cocaine.

The effect of 'binge' cocaine administration on the expression of cyclin-dependent kinase 5 and its activator p35 in various regions of rat brain.

The present study was aimed at determining whether the administration of cocaine in 'binge' pattern regimen that evoked tolerance to the locomotor stimulant effects of cocaine also influenced the expression of cyclin-dependent kinase 5 (Cdk5) and its activator p35 in the amygdala, medial prefrontal cortex, nucleus accumbens septi and caudate-putamen. Western blot techniques revealed that acute and repeated 'binge' cocaine decreased expression of the Cdk5 protein in the amygdala. In the medial prefrontal cortex, only exposure to repeated 'binge' cocaine decreased the content of the Cdk5 protein. 'Binge' cocaine administration also altered the expression of Cdk5 activator p35 protein. In the amygdala, only repeated 'binge' cocaine decreased the expression of p35, while in the medial prefrontal cortex, a decrease was observed after acute and repeated 'binge' cocaine exposure. In neither the nucleus accumbens septi nor the caudate-putamen acute or repeated 'binge' cocaine modified the expression of Cdk5 and p35. The above data indicate that in contrast to sensitizing doses of cocaine, a single and repeated binge of cocaine, which evoked tolerance to its locomotor stimulant effects, decreases expression of Cdk5 and p35 and possibly decreases the efficacy of neurotransmission or induces brain plastic changes regulated by Cdk5 and its activator p35.