Increase in Alzheimer's related markers preceeds memory disturbances: studies in vasopressin-deficient Brattleboro rat.

Alzheimer's disease (AD) is the most common form of dementia in the elderly. For more effective therapy early diagnostic markers could be beneficial. Therefore we compared one year old rats with adults and examined if changes in possible brain markers of AD preceeded memory decline. We also tested if vasopressin-deficient animals were useful model of AD as vasopressin has well known positive effect on memory and AD patient has decreased vasopressin production. We compared adult (3 month) and old (12 month), normal and vasopressin-deficient Brattleboro rats. To receive a comprehensive picture about their memory we examined their social discrimination, object discrimination and conditioned learning abilities (shuttle box). Amyloid precursor protein (APP), mitogen-activated protein kinase 1 (MAPK1), ß-actin and tryptophan 2,3-dioxygenase 2 (TDO2) mRNA levels was measured by quantitative PCR. There was no difference between the memory of adult and aged groups. The vasopressin-deficient rats at both ages showed a weaker performance in the course of social and object discrimination tests and a higher escape failure during the shuttle box experiment. The brain marker mRNAs of the elder animals were higher than the levels of the adults, but the absence of vasopressin had no influence on them. Thus, the one year old rats showed elevated levels of AD-related markers, but memory deficits were observable only in vasopressin deficient animals. Vasopressin does not seem to have pathogenic role in AD. Changes in the studied markers might predict later symptoms, although further studies are required for confirmation.

Blunted HPA axis response in lactating, vasopressin-deficient Brattleboro rats.

Adaptation to stress is a basic phenomenon in mammalian life that is mandatorily associated with the activity of the hypothalamic-pituitary-adrenal (HPA) axis. An increased resting activity of the HPA axis can be measured during pregnancy and lactation, suggesting that these reproductive states lead to chronic load in females. In this study, we examined the consequences of the congenital lack of vasopressin on the activity of the HPA axis during lactation using vasopressin-deficient Brattleboro rats. Virgin and lactating, homozygous vasopressin-deficient rats were compared with control, heterozygous rats. In control dams compared with virgins, physiological changes similar to those observed in a chronic stress state (thymus involution, adrenal gland hyperplasia, elevation of proopiomelanocortin mRNA levels in the adenohypophysis, and resting plasma corticosterone levels) were observed. In vasopressin-deficient dams, adrenal gland hyperplasia and resting corticosterone level elevations were not observed. Corticotropin-releasing hormone (Crh) mRNA levels in the hypothalamic paraventricular nucleus were elevated in only the control dams, while oxytocin (OT) mRNA levels were higher in vasopressin-deficient virgins and lactation induced a further increase in both the genotypes. Suckling-induced ACTH and corticosterone level elevations were blunted in vasopressin-deficient dams. Anaphylactoid reaction (i.v. egg white) and insulin-induced hypoglycemia stimulated the HPA axis, which were blunted in lactating rats compared with the virgins and in vasopressin-deficient rats compared with the controls without interaction of the two factors. Vasopressin seems to contribute to the physiological changes observed during lactation mimicking a chronic stress state, but its role in acute HPA axis regulation during lactation seems to be similar to that observed in virgins. If vasopressin is congenitally absent, OT, but not the CRH, compensates for the missing vasopressin; however, the functional restitution remains incomplete.

Do stress hormones connect environmental effects with behavior in the forced swim test?

Forced swim test (FST) is a widely used test for antidepressant development. Depression is a stress related disease, as hormones of the stress-axis can modify mood. However it is not clear, how the appearance of depressive-like behavior (floating) in FST is connected with changes in the stress-hormone levels. We hypothesized, that different manipulations would alter the behavior through changes in stress-hormone levels. First the effect of environmental alterations was studied. Increasing water-temperature enhanced floating time together with a decrease in adrenocorticotropin levels. During the dark phase of the day rats spent more time with floating independently from the actual lighting. Neither the phase nor the actual lighting had significant effect on adrenocorticotropin concentrations with higher corticosterone levels during the dark phase. At greater water depth rats float less but the size of animals had no effect. Water depth did not influence adrenocorticotropin and corticosterone responses, but the size of the rats significantly affected both factors. Secondly, administration of imipramine reduced floating and adrenocorticotropin level without affecting corticosterone. Despite the known connection between depression and stress we did not find a correlation between floating behavior and hormone levels. As an alternative mechanism imipramine-induced heart rate and core body temperature decrease was found by telemetric approach. This study is the first summary in rats examining the effect of wide range of environmental alterations during FST. It seems likely that both brain monoamines and stress-axis take part in the development of depression, but these pathways are regulated independently.

Lack of vasopressin does not prevent the behavioural and endocrine changes induced by chronic unpredictable stress.

Vasopressin (VP) plays an important role in hypothalamo-pituitary-adrenal (HPA) axis regulation and in stress-related disorders. Our previous studies confirmed the role of VP in acute situations, where VP-deficient Brattleboro rats had less depression-like behaviour compared to animals that express VP. In this study, we test the hypothesis that VP-deficient rats are more resistant to the development of chronic HPA axis hyperactivity and depression-like symptoms after chronic unpredictable stress (CUS). Male VP-deficient Brattleboro rats were compared to their heterozygous littermates (controls). CUS consisted of different mild stimuli for 5 weeks. Elevated plus maze and forced swim test were used for behavioural characterization, while organs and blood for HPA axis parameters were collected at the end of the experiment. In controls, CUS resulted in the development of chronic stress state characterized by typical somatic (body weight reduction, thymus involution) and endocrine changes (resting plasma ACTH and corticosterone elevation and POMC mRNA elevation in anterior lobe of the pituitary). Floating time in the forced swim test was enhanced together with reduced open arm entries on elevated plus maze and a reduction in daily food intake. Unexpectedly, the lack of VP did not alter the effect of CUS on the somatic and behavioural measures, but only prevented CUS-induced corticosterone changes. In conclusion, lifelong VP-deficiency has a positive effect on corticosterone elevation following CUS but does not affect the behavioural consequences of CUS. It is likely that the interplay of several related factors, rather than an alteration in a single neuropeptide, modulates behaviour and disease pathogenesis.

[The effects of duloxetine on beta-actin stress response in rat brain]. / A duloxetin hatása a beta-aktin stresszválaszra patkány agyban.

Depression is a frequent prodromal symptom of Alzheimer's disease (AD). Stress factors play an important role in the etiopathology of both diseases, since increased corticosteroid levels caused by chronic stress indirectly induce neuronal damage. The aim of our experiments was to evaluate the changes induced by stress in the transcription of amyloid precursor protein (APP), mitogen activated protein kinase-1 (MAPK-1) and beta-actin, of which the latest plays a leading role in synaptic plasticity. Additionally we intended to examine how duloxetine - a serotonin-norepinephrin reuptake inhibitor type antidepressant - would modify the stress-induced changes. Wistar rats were exposed to immobilization stress for five hours daily through 21 days, while part of the animals received 45 mg/bwkg of duloxetine. At the end of the third week total RNA was purified from the cortex and hippocampus. The amount of beta-actin, APP and MAPK-1 mRNA was determined by real time PCR method. On protein level, semiquantitative measurement was performed by Western blot. The expression of beta-actin mRNA in the animals exposed to stress was four times as intense as in the control group. The increase in the beta-actin mRNA levels was repressed by the duloxetine treatment. In the case of APP and MAPK-1 no changes were detected. According to the Western blot results, the antidepressant treatment slightly, the drug along with the stress treatment strongly decreased the amount of the beta-actin protein. Our findings indicate that antidepressant treatment with duloxetine could play a protective role against the chronic stress-induced changes in the nervous system, such as disorders of synaptic plasticity, and the consequent cognitive dysfunctions in case of both affective disorders and AD.

Intermittent prenatal MDMA exposure alters physiological but not mood related parameters in adult rat offspring.

The recreational party drug "ecstasy" (3,4-methylenedioxymethamphetamine MDMA) is particularly popular among young adults who are in the childbearing age and thus there is a substantial risk of prenatal MDMA exposure. We applied an intermittent treatment protocol with an early first injection on pregnant Wistar rats (15 mg/kg MDMA s.c. on the E4, E11 and E18 days of gestation) to examine the potential physiological, endocrine and behavioral effects on adult male and female offspring. Prenatal MDMA-treatment provoked reduced body weight of offspring from the birth as far as the adulthood. Adult MDMA-offspring had a reduced blood-glucose concentration and hematocrit, altered relative spleen and thymus weight, had lower performance on wire suspension test and on the first trial of rotarod test. In contrast, no alteration in the locomotor activity was found. Anxiety and depression related behavioral parameters in elevated plus maze, sucrose preference or forced swimming tests were normal. MDMA-offspring had elevated concentration of the ACTH-precursor proopiomelanocortin and male MDMA-offspring exhibited elevated blood corticosterone concentration. No significant alteration was detected in the serotonergic marker tryptophan-hydroxylase and the catcholaminergic marker tyrosine-hydroxylase immunoreactive fiber densities in MDMA-offspring. The mothers exhibited reduced densities of serotonergic but not catecholaminergic fibers after the MDMA treatment. Our findings suggest that an intermittent prenatal MDMA exposure with an early first injection and a relatively low cumulative dose provokes mild but significant alterations in physical-physiological parameters and reduces motor skill learning in adulthood. In contrast, these adult offspring do not produce anxiety or depression like behavior.

[The transcription of the amyloid precursor protein and tryptophan 2,3-dioxygenase genes are increased by aging in the rat brain]. / Az öregedés fokozza az amiloid prekurzor protein és a triptofán 2,3-dioxigenáz gének transzkripcióját patkányagyban.

Aging itself is considered as a major risk factor of dementia. The prevalence of the Alzheimer's disease (AD) is increasing exponentially after the age of 65 and doubles every 5 years. The major aim of our present research was to examine the effect of aging on the transcription of certain genes associated with neurodegenerative disorders in the rat brain. The influence of the vasopressin (VP) hormone was also examined in the same experimental paradigm. Age dependent transcriptional changes of the following four genes were examined in the cerebral cortex: the first was the gene of the amyloid precursor protein (APP) which is abnormally cleaved to toxic beta-amyloid fragments. These aggregated peptides are the major components of the senile plaques in the AD brain. The second one was the mitogen-activated protein kinase (MAPK1) gene. The MAPK is involved in the abnormal hyperphosphorylation of the tau-protein which results in aggregated neurofibrillary tangles. The beta-actin gene was the third one. The protein product of this gene is considered to be involved in synaptogenesis, neuronal plasticity and clinical conditions like depression and AD. The last one was the gene of the tryptophan 2,3-dioxygenase (TDO2) enzyme. The activity of this enzyme is considered as a rate limiting factor in the metabolism of the neuro-immune modulator quinolinic acid (QUIN). The transciptional activity of young (2.5 months) and aged (13 months) Brattleboro rats with or without VP expression were compared by means of real time PCR technique. The cortical transciptional activity of the APP and TDO2 genes were increased in the aged animals as compared with the activity of the young ones, and this effect was independent on the presence of the VP. Our results indicate the importance of certain age dependent transcriptional changes might influence the mechanism of AD and other neurodegenerative disorders.

The stimuli-specific role of vasopressin in the hypothalamus-pituitary-adrenal axis response to stress.

Adaptation to a constantly changing environment is fundamental to every living organism. The hypothalamic-pituitary-adrenocortical (HPA) axis is a key component of the adaptation process. The present study tests the hypothesis that vasopressin (AVP) is required for the HPA response to acute stimuli. To accomplish this, naturally AVP-deficient Brattleboro rats were exposed to a wide range of stimuli and their HPA response was compared with heterozygous littermates. The circadian rhythmicity of plasma ACTH and corticosterone was not different between the two genotypes. The ACTH and corticosterone response to volume load, restraint or aggressive attack were decreased in AVP-deficient rats. The stress-induced increase in ACTH, but not corticosterone, was significantly impaired in AVP-deficient animals after novelty, elevated plus-maze, forced swim, hypoglycaemia, ulcerogenic cold immobilisation, lipopolysaccharide, hypertonic saline and egg white injection. The HPA response to social avoidance, ether inhalation and footshock was not different between the genotypes. In vitro, the hypophysis of AVP-deficient animals showed a reduction in stimulated ACTH production and their adrenal glands were hyporeactive to ACTH. A dissociation between the ACTH and corticosterone response was observed in several experiments and could not be explained by an earlier ACTH peak or enhanced adrenal sensitivity, suggesting the existence of paraadenohypophyseal neuroendocrine regulators. Loss of AVP affected the HPA response to a wide variety of stressors. Interestingly, the contribution of AVP to the HPA response was not specific for, nor limited to, a known stressor category. Thus, there is a context-specific requirement for AVP in stress-induced activation of the HPA axis.

Capsaicin promotes the amyloidogenic route of brain amyloid precursor protein processing.

Besides being an important component of spices used worldwide, capsaicin has wide-ranging therapeutic potential as a hypolipidemic, antioxidant and anti-inflammatory agent. Accordingly, it is very important to investigate the long-term effect of capsaicin in the pathogenesis of Alzheimer's disease. In this study, the effects of capsaicin on the processing of amyloid precursor protein (APP) were investigated in an in vivo model. The APP mRNA and protein levels were examined in the brain cortices of control and capsaicin-treated rats. The protein kinase C (PKC) translocation state in the soluble and membrane-bound fractions and the levels of beta-secretase (BACE) were also evaluated. Capsaicin enhanced the level of membrane-bound APP 1.7-fold. The APP mRNA and PKC and BACE protein levels were unchanged after capsaicin treatment. These in vivo data indicate that capsaicin is able to interfere with the brain APP metabolism by promoting the amyloidogenic route. We suggest that PKC is not involved in the mechanism underlying the effects.

Vasopressin administration into the paraventricular nucleus normalizes plasma oxytocin and corticosterone levels in Brattleboro rats.

Adult male rats of the Brattleboro strain were used to investigate the impact of the congenital absence of vasopressin on plasma adrenocorticotropin, corticosterone, and oxytocin concentrations as well as the release pattern of oxytocin within the hypothalamic paraventricular nucleus (PVN), in response to a 10-min forced swimming session. Measurement of adrenocorticotropin in plasma samples collected via chronically implanted jugular venous catheters revealed virtually identical stress responses for vasopressin-lacking Brattleboro (KO) and intact control animals. In contrast, plasma corticosterone and oxytocin levels were found to be significantly elevated 105 min after onset of the stressor in KO animals only. Microdialysis samples collected from the extracellular fluid of the PVN showed significantly higher levels of oxytocin both under basal conditions and in response to stressor exposure in KO vs. intact control animals accompanied by elevated oxytocin mRNA levels in the PVN of KO rats. These findings suggest that the increased oxytocin levels in the PVN caused by the congenital absence of vasopressin may contribute to normal adrenocorticotropin stress responses in KO animals. However, whereas the stressor-induced elevation of plasma oxytocin in KO rats may be responsible for their maintained corticosterone levels, oxytocin seems unable to fully compensate for the lack of vasopressin. This hypothesis was tested by retrodialyzing synthetic vasopressin into the PVN area concomitantly with blood sampling in KO animals. Indeed, this treatment normalized plasma oxytocin and corticosterone levels 105 min after forced swimming. Thus, endogenous vasopressin released within the PVN is likely to act as a paracrine signal to facilitate the return of plasma oxytocin and corticosterone to basal levels after acute stressor exposure.

Control of the hypothalamo-pituitary-adrenal axis in the neonatal period: adrenocorticotropin and corticosterone stress responses dissociate in vasopressin-deficient brattleboro rats.

In adulthood the hypothalamo-pituitary-adrenal axis is controlled by both CRH and arginine vasopressin (AVP). However, in neonates CRH secretion is very low, whereas AVP secretion is fully functional. This suggests that the role of AVP is more pronounced in young than in adult rats. We investigated the role of AVP by studying stress responses in 5, 10, and 20-d-old AVP-deficient Brattleboro rats. Two different stressors were applied: 24-h maternal separation and Hypnorm Grove Oxford UK injections. In heterozygous controls (that do express AVP), both stressors increased plasma ACTH and corticosterone. The ACTH stress response disappeared in AVP-deficient rats, demonstrating that during the perinatal period, the secretion of this hormone is controlled by AVP. Surprisingly, corticosterone responses remained intact in AVP-deficient rats. Similar findings were obtained after 1-, 4-, 12-, and 24-h long maternal separations. Thus, preserved corticosterone stress responses were not explained by changes in the timing of ACTH secretion. In vitro experiments suggested that the dissociation of ACTH and corticosterone stress responses can only be partly explained by higher ACTH responsiveness of the adrenal cortex in AVP-deficient rats. Together, our results show that in neonatal periods, AVP is crucial for the expression of ACTH stress responses, but neither AVP nor ACTH is necessary for the induction of corticosterone stress responses. Discrepant ACTH and corticosterone stress responses may reflect compensatory mechanisms activated by AVP deficiency, but disparate findings suggest that they rather depict a neonate-specific mechanism of hypothalamo-pituitary-adrenal-axis control.

Congenital vasopressin deficiency and acute and chronic opiate effects on hypothalamo-pituitary-adrenal axis activity in Brattleboro rats.

A growing body of evidence suggests that vasopressinergic activity in the hypothalamus is important in stress-related behaviors (like drug abuse) in line with a role in the regulation of the hypothalamo-pituitary-adrenal axis (HPA). We hypothesized that in the naturally vasopressin-deficient Brattleboro rat, acute and chronic morphine treatment may lead to reduced HPA axis activity. Rats were treated either with a single dose of morphine (10 mg/kg subcutaneously) and serial blood samples were taken or were treated twice daily with increasing doses of morphine (10-100 mg/kg subcutaneously) for 16 days and animals were killed by decapitation 4 or 16 h after the last injection. Single morphine injection induced a biphasic ACTH and corticosterone elevation with smaller increases in vasopressin-deficient rats. Chronic morphine treatment induced the typical somatic and HPA axis changes of chronic stress; the absence of vasopressin did not prevent these changes. In rats repeatedly treated with morphine plasma, ACTH and corticosterone levels were elevated both 4 and 16 h after the last injection (short and long withdrawal) and the absence of vasopressin attenuated this response. Our data suggest that vasopressin plays a prominent role in morphine treatment and withdrawal-induced acute hormonal changes, but does not affect development of chronic hyperactivity of the HPA axis.

Gender-specific regulation of the hypothalamo-pituitary-adrenal axis and the role of vasopressin during the neonatal period.

Studies in arginine vasopressin (AVP)-deficient Brattleboro rats suggest that AVP is the predominant secretagogue during the perinatal period. Here we tested the hypothesis that congenital lack of vasopressin differentially modifies the stress reactivity of male and female rat pups. Vasopressin-producing (heterozygous, AVP+) and AVP-deficient (AVP-) Brattleboro rat pups of both genders were used. In 10-day-old pups, 24-h maternal separation and single, as well as repeated, ether inhalation induced remarkable adrenocorticotropin (ACTH) elevation only in AVP+ pups, supporting the role of vasopressin in hypothalamo-pituitary-adrenal (HPA) axis regulation. Surprisingly, the corticosterone elevations were even more pronounced in AVP- pups, suggesting the possibility of an ACTH-independent corticosterone-secretion regulation. In the case of maternal separation, both the plasma ACTH and corticosterone levels were higher in females than in males, while in case of ether inhalation only the ACTH levels were higher in females. Gender did not influence the stress reactivity or the effect of the genotype. We conclude that the gender of the pups did not profoundly influence HPA axis activity (the mechanism seems to be the same), but in contrast to the general view, we suggest that the females have a more active HPA axis than the males already during the neonatal period. However, the resting corticosterone elevation-well known in adult females- is missing.

The role of vasopressin in chronic stress studied in a chronic mild stress model of depression.

BACKGROUND AND PURPOSE: Vasopressin plays an important role in the hypothalamo-pituitary-adrenal axis regulation as well as in stress-related disorders. A common view suggested that the role of vasopressin is especially important during chronic stresses. Here we tested the hypothesis that vasopressin-deficient rats may be more resistant to the development of chronic hypothalamo-pituitary-adrenal axis hyperactivity after chronic mild stress. METHODS: Male vasopressin deficient Brattleboro rats were compared to their heterozygous litter mates. Chronic mild stress consisted of different mild stimuli (e.g. wet cages, restraint) for 6 week. The corticosterone changes were followed by repeated tail cutting and organs and blood were collected from decapitated rats. RESULTS: In controls, chronic mild stress resulted in symptoms of chronic stress state characterized by typical somatic (body weight reduction, thymus involution) and endocrine changes (resting plasma ACTH and corticosterone elevation and POMC mRNA elevation in anterior lobe of the pituitary). Unexpectedly, the lack of vasopressin could not influence any chronic mild stress-induced changes. CONCLUSION: Somatic changes and endocrine effects of chronic mild stress are similar in control and vasopressin deficient animals. This suggests that either vasopressin is not indispensable for activating the hypothalamo-pituitary-adrenal axis by chronic stress or the absence of vasopressin is compensated by other mediators (e.g. CRH) in Brattleboro rats.