Vitamin K Deficiency Induced by Warfarin Is Associated With Cognitive and Behavioral Perturbations, and Alterations in Brain Sphingolipids in Rats.

Initially discovered for its role in blood coagulation, there is now convincing evidence that vitamin K (VK) has important actions in the nervous system. In brain, VK is present in the form of menaquinone-4 (MK-4), a byproduct of the main dietary source, phylloquinone. It contributes to the biological activation of various proteins (i.e., Gas6), and participates in the synthesis of sphingolipids, a class of lipids widely present in brain cell membranes with important cell signaling functions. In a previous study, we reported that lifetime consumption of a low VK diet resulted in mild cognitive impairment in aged rats, a finding associated with an alteration of the sphingolipid profile. To confirm the role of VK as it relates to sphingolipids, cognition, and behavior outside the context of aging, we conducted a study of acute VK deficiency using a pharmacological model of VK deficiency in brain. In this procedure, rats (8 weeks) are maintained on a ratio of warfarin (a VK antagonist) to VK whereby coagulation is maintained while inducing VK deficiency in extrahepatic tissues. After 10 weeks of treatment, rats who were subjected to the warfarin plus phylloquinone protocol (WVK) exhibited longer latencies in the Morris water maze test as well as lower locomotor activity and exploratory behavior in the open field test, when compared to control rats. The WVK treatment resulted in a dramatic decrease in MK-4 level in all brain regions despite the presence of high local concentrations of phylloquinone, which suggests an inhibition of the biosynthetic MK-4 pathway in the presence of warfarin. Additionally, WVK treatment affected sphingolipid concentrations in key brain regions, notably those of the ganglioside family. Finally, brain MK-4 was correlated with performances in the open field test. This study confirms the modulatory role of VK in cognition and behavior and the implication of sphingolipids, notably those of the ganglioside family.

Absence of neurological abnormalities in mice homozygous for the Polr3a G672E hypomyelinating leukodystrophy mutation.

Recessive mutations in the ubiquitously expressed POLR3A gene cause one of the most frequent forms of childhood-onset hypomyelinating leukodystrophy (HLD): POLR3-HLD. POLR3A encodes the largest subunit of RNA Polymerase III (Pol III), which is responsible for the transcription of transfer RNAs (tRNAs) and a large array of other small non-coding RNAs. In order to study the central nervous system pathophysiology of the disease, we introduced the French Canadian founder Polr3a mutation c.2015G > A (p.G672E) in mice, generating homozygous knock-in (KI/KI) as well as compound heterozygous mice for one Polr3a KI and one null allele (KI/KO). Both KI/KI and KI/KO mice are viable and are able to reproduce. To establish if they manifest a motor phenotype, WT, KI/KI and KI/KO mice were submitted to a battery of behavioral tests over one year. The KI/KI and KI/KO mice have overall normal balance, muscle strength and general locomotion. Cerebral and cerebellar Luxol Fast Blue staining and measurement of levels of myelin proteins showed no significant differences between the three groups, suggesting that myelination is not overtly impaired in Polr3a KI/KI and KI/KO mice. Finally, expression levels of several Pol III transcripts in the brain showed no statistically significant differences. We conclude that the first transgenic mice with a leukodystrophy-causing Polr3a mutation do not recapitulate the childhood-onset HLD observed in the majority of human patients with POLR3A mutations, and provide essential information to guide selection of Polr3a mutations for developing future mouse models of the disease.

FXR1P limits long-term memory, long-lasting synaptic potentiation, and de novo GluA2 translation.

Translational control of mRNAs allows for rapid and selective changes in synaptic protein expression that are required for long-lasting plasticity and memory formation in the brain. Fragile X Related Protein 1 (FXR1P) is an RNA-binding protein that controls mRNA translation in nonneuronal cells and colocalizes with translational machinery in neurons. However, its neuronal mRNA targets and role in the brain are unknown. Here, we demonstrate that removal of FXR1P from the forebrain of postnatal mice selectively enhances long-term storage of spatial memories, hippocampal late-phase long-term potentiation (L-LTP), and de novo GluA2 synthesis. Furthermore, FXR1P binds specifically to the 5' UTR of GluA2 mRNA to repress translation and limit the amount of GluA2 that is incorporated at potentiated synapses. This study uncovers a mechanism for regulating long-lasting synaptic plasticity and spatial memory formation and reveals an unexpected divergent role of FXR1P among Fragile X proteins in brain plasticity.

A controlled evaluation of a targeted early case detection intervention for reducing delay in treatment of first episode psychosis.

PURPOSE: Interventions aimed at influencing specific pathways to care may reduce the duration of untreated psychosis (DUP). The purpose of this study was to reduce delay in referral to an early intervention service following first contact for help seeking for psychosis in a defined urban catchment area. METHODS: Using a historical control design, we conducted a targeted intervention comprised of intensive training and education regarding first onset of psychosis and benefits of early intervention with all potential points of contact in pathways to care in a defined catchment area of a specialized early intervention (EI) service. Data collected on different components of DUP [help seeking (H-DUP) and referral (R-DUP)] and demographic and clinical characteristics on patients seeking treatment of first episode of a psychotic (FEP) disorder accepted in the EI service for 3 years before and 3 years after the intervention were contrasted. No other systemic changes occurred in this catchment area during this period. RESULTS: There was a significant increase in the number of cases of FEP referred to the EI service post-intervention from hospitals other than the parent institute, in the proportion with a diagnosis of affective psychosis and proportion of patients from a lower socioeconomic status (SES). Although largest proportion of patients continued to make their first contact with community services, the latter failed to refer cases of psychosis to the EI service despite making a large number of mostly non-cases (for psychosis). The intervention had no direct effect on total DUP, R-DUP or H-DUP. After controlling for diagnosis and SES, post-intervention, R-DUP remained significantly lower for patients making first contact with the emergency service of the parent institute. CONCLUSIONS: The intervention failed to reduce R-DUP for patients making contact with or referred from community services and most patients still used hospital resources to enter treatment at the EI service.

Pioglitazone improves reversal learning and exerts mixed cerebrovascular effects in a mouse model of Alzheimer's disease with combined amyloid-ß and cerebrovascular pathology.

Animal models of Alzheimer's disease (AD) are invaluable in dissecting the pathogenic mechanisms and assessing the efficacy of potential new therapies. Here, we used the peroxisome proliferator-activated receptor gamma agonist pioglitazone in an attempt to rescue the pathogenic phenotype in adult (12 months) and aged (>18 months) bitransgenic A/T mice that overexpress a mutated human amyloid precursor protein (APPSwe,Ind) and a constitutively active form of transforming growth factor-ß1 (TGF-ß1). A/T mice recapitulate the AD-related cognitive deficits, amyloid beta (Aß) and cerebrovascular pathologies, as well as the altered metabolic and vascular coupling responses to increased neuronal activity. Pioglitazone normalized neurometabolic and neurovascular coupling responses to sensory stimulation, and reduced cortical astroglial and hippocampal microglial activation in both age groups. Spatial learning and memory deficits in the Morris water maze were not rescued by pioglitazone, but reversal learning was improved in the adult cohort notwithstanding a progressing Aß pathology. While pioglitazone preserved the constitutive nitric oxide synthesis in the vessel wall, it unexpectedly failed to restore cerebrovascular reactivity in A/T mice and even exacerbated the dilatory deficits. These data demonstrate pioglitazone's efficacy on selective AD hallmarks in a complex AD mouse model of comorbid amyloidosis and cerebrovascular pathology. They further suggest a potential benefit of pioglitazone in managing neuroinflammation, cerebral perfusion and glucose metabolism in AD patients devoid of cerebrovascular pathology.

Lifelong low-phylloquinone intake is associated with cognitive impairments in old rats.

In a previous report, we showed vitamin K to preferentially accumulate in brain regions rich in white matter and to positively correlate with certain sphingolipids. In rodents, pharmacological vitamin K deficiency has resulted in behavioral perturbations. To gain insight on the role of vitamin K status on brain function, we investigated learning abilities (Morris water maze), motor activity (open field), and anxiety (elevated plus maze) in distinct groups of 6-, 12-, and 20-mo-old female Sprague-Dawley rats that had been fed diets containing low (L; ~80 µg/kg diet), adequate (A; ~500 µg/kg diet), or high (H; ~2000 µg/kg diet) levels of phylloquinone (µg/kg diet; n = 9-12/diet) since weaning. In 20-mo-old rats, sphingolipids (cerebroside, sulfatide, sphingomyelin, ceramide, and gangliosides), phylloquinone, and menaquinone-4 were also assessed in cerebellum, midbrain, pons medulla, striatum, and hippocampus. Lifetime consumption of a low-vitamin K diet resulted in cognitive deficits in the 20-mo-old rats, with those in the L group having longer latencies than those in the H group (P < 0.05); this was associated with higher concentrations of ceramides in the hippocampus (P < 0.05) and lower gangliosides in the pons medulla and midbrain (P < 0.05). The low-vitamin K diet did not affect cognition at 6 and 12 mo of age, nor did it affect motor activity or anxiety at any age. Although much remains to be elucidated about the mechanism of action of vitamin K in cognition, this report points to vitamin K as an important nutritional factor contributing to cognitive health during aging.

Long term zoledronic acid during androgen blockade for prostate cancer.

OBJECTIVES: To evaluate the effect of zoledronic acid on androgen deprivation therapy in patients with hormone-sensitive prostate cancer by measuring the percentage change in lumbar-spine bone mineral density (BMD) at 12 and 24 months. MATERIALS AND METHODS: An open-label, multicenter, randomized, two-phase study was conducted in patients with hormone-sensitive prostate cancer (N = 200) receiving 10.8 mg goserelin acetate with or without zoledronic acid (4 mg intravenously) every 3 months. In phase I, patients were randomized to goserelin acetate alone or goserelin acetate plus zoledronic acid for 12 months. In phase II, patients receiving goserelin acetate plus zoledronic acid continued treatment for up to a total of 24 months, whereas patients receiving goserelin acetate alone were randomized to goserelin acetate alone or goserelin acetate plus zoledronic acid for an additional 12 months. Lumbar-spine, femoral-neck, and total-hip BMD were assessed at 6, 12, and 24 months. Additional assessments included height change, laboratory studies, bone scans, radiographs, and computed tomography scans. RESULTS: Significant BMD differences between patients receiving goserelin acetate alone and goserelin acetate plus zoledronic acid were observed at the 12-month (p

Naturally occurring variations in maternal care modulate the effects of repeated neonatal pain on behavioral sensitivity to thermal pain in the adult offspring.

Repeated pain during a critical period of development can have long-term behavioral and physiological consequences in both human and animals. We previously showed that rat mothers caring for pups subjected to mild pain in neonatal life increased pup licking and grooming behavior. Therefore, we tested whether naturally occurring variations in maternal behavior would modulate the effects of repeated mild inflammatory pain on behavioral responses to pain and stress in the adult male offspring. Rat pups were either uninjected (UI) or injected twice daily between PND3 and PND14 with either saline (0.9%) or formalin (0.2-0.4%) in the footpad of the hindpaw. Maternal behavior (pup licking and grooming) was recorded under basal conditions and after reunion with the litter post injection to determine maternal phenotype (High, Middle, Low licking). Adult offspring (PND60) were tested for their thermal sensitivity, inflammatory pain responses after formalin injection and neuroendocrine responses to formalin injection. Maternal phenotype significantly altered pain sensitivity after thermal stimulation, but not formalin injection. Offspring from the High licking mothers displayed increased withdrawal latencies compared to offspring from Low mothers, regardless of neonatal treatment. Pain responses after formalin injection were higher in offspring receiving formalin as neonates compared to saline-treated or uninjected rats, demonstrating a long lasting increased sensitivity to inflammatory pain. Neuroendocrine responses to pain stress were not affected by neonatal treatment. These data suggest that changes in maternal behavior can influence some modalities of pain sensitivity and that repeated mild inflammatory pain in neonatal period causes hypersensitivity to formalin in the adult offspring.

Antidepressant action of agomelatine (S 20098) in a transgenic mouse model.

The aim of this study was to evaluate the efficacy of agomelatine (S 20098) to accelerate reversal of the neuroendocrinological, behavioural and cyclical changes seen in a transgenic mouse model of the neuroendocrine characteristics of depression. The effects of agomelatine were assessed in transgenic mice with low glucocorticoid receptor (GR) function, after acute stress or induced phase shift, and compared to desipramine and melatonin. Mice were injected 2 h before the onset of the dark period with agomelatine (10 mg/kg, i.p.), desipramine (10 mg/kg, i.p.), melatonin (10 mg/kg, i.p.) or vehicle (hydroxy-ethyl-cellulose (HEC) 1%) each day for 21 to 42 days. Agomelatine was effective in reversing the transgenic mouse behavioural changes noted in the Porsolt forced swim test as well as in the elevated plus maze. Both the number of open arm entries and the total time spent in open arms of the elevated plus maze is greatly increased in transgenic mice. The mean time spent in open arms is exquisitely sensitive to reversal by agomelatine and desipramine. Agomelatine also markedly accelerated readjustment of circadian cycles of temperature and activity following an induced phase shift. This action of agomelatine was superior to that of melatonin while desipramine was without effect. The accelerating effect of agomelatine was particularly notable if treatment was started 3 weeks prior to the induced phase shift. Agomelatine treatment did not cause any major change in corticosterone or adrenocorticotropic hormone (ACTH) concentrations nor in vasopressin (AVP), corticotropin-releasing hormone (CRH), GR and mineralocorticoid receptor (MR) mRNAs levels, which make it unlikely that the mechanism of agomelatine action is related to hypothalamic-pituitary-adrenocortical (HPA) axis changes. The present study shows that agomelatine displays some characteristics of antidepressant drug action in the transgenic mouse model, effects that could be partially related to its chronobiotic properties.

Antidepressant-like effects of neurokinin receptor antagonists in the forced swim test in the rat.

Although a wide assortment of agents is currently available for the treatment of depression, this disorder remains poorly managed in a large proportion of patients. Traditional antidepressant treatments target the biogenic amine systems. However, a growing body of evidence is implicating the involvement of neuropeptides in depression, especially the neurokinin substance P. This study evaluated the effects of selective antagonists of the tachykinin NK1, NK2, and NK3 receptors in the forced swim test, a commonly used screen for antidepressants. Rats were given CP-96,345 (2S, 3S)-cis-2-(diphenylmethyl)-N-[(2-methoxyphenyl)-methyl]-1-azabicyclo[2.2.2]octan-3-amine, SR 48968 (S)-N-methyl-N[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl)-butyl]benzamide, or SR 142801 (S)-(N)-(1-(3-(1-benzoyl-3-(3,4-dichlorophenyl) piperidin-3-yl) propyl)-4-phenylpiperidin-4-yl)-N-methylacetamide, antagonists of the NK1, NK2, and NK3 receptors, respectively, at doses of 2.5, 5, and 10 mg/kg, intraperitoneally (i.p.). The time of immobility during the forced swim test was used as an indicator of antidepressant activity of the antagonists. All antagonists decreased immobility times. CP-96,345 and SR 142801 showed dose-related effects; SR 48968 had its maximum effect at 2.5 mg/kg. The magnitude of the effects of the neurokinin receptor antagonists was approximately the same as that of amitriptyline and desipramine, two traditional antidepressants, both given at 10 mg/kg, i.p. This study provides comparative data on the relative effectiveness of NK1, NK2, and NK3 receptor antagonists in this screen for antidepressant drug activity.

Menaquinone-4 concentration is correlated with sphingolipid concentrations in rat brain.

Studies with animals support a role for vitamin K (VK) in the biosynthesis of sphingolipids, a class of complex lipids present in high concentrations in the brain. In mice and rats, VK deficiency decreases levels of brain sulfatides and causes behavioral alterations. In light of its heterogeneity and to better understand the role of VK in the brain, we characterized the distribution of the two main VK vitamers, phylloquinone (K1) and menaquinone-4 (MK-4), in nine distinct brain regions. Weaning female Sprague-Dawley rats (n=5/dietary group) were fed diets containing either low (L, 80 microg/kg diet), adequate (A, 500 microg/kg diet) or high (H, 2000 microg/kg diet) levels of K1 for 6 mo. The main form of VK in the brain was MK-4, and it was present in significantly higher concentrations in myelinated regions (the pons medulla and midbrain) than in nonmyelinated regions. Both regional K1 and MK-4 increased with K1 intake (P<0.05). Sphingolipid distribution varied across brain regions (P<0.001) but was not affected by K1 intake. In the L and A groups but not the H group, brain MK-4 concentration was positively correlated with the concentrations of sulfatides (L, r=0.518; A, r=0.479) and sphingomyelin (L, r=0.515; A, r=0.426), and negatively correlated with ganglioside concentration (L, r=-0.398); A, r=-0.353). Sphingolipids are involved in major cellular events such as cell proliferation, differentiation and survival. The strong associations reported here between brain MK-4 and sphingomyelin, sulfatides and gangliosides suggest that this vitamer may play an important role in the brain.

Effects of ovariectomy and estradiol on acoustic startle responses in rats.

Long-term (3 months) ovariectomized (OVX) rats were used to model hormone withdrawal as occurring in menopause. We previously reported alterations in brain dopamine (DA), GABA and serotonin receptors following ovariectomy in this model. To assess the functional effect of these biochemical changes, we compared rats that were intact, OVX and OVX-treated with 17beta-estradiol (E(2); OVX+E(2)) for 2 weeks on measures of their acoustic startle responses (ASR) and prepulse inhibition (PPI) of acoustic startle. The effects of a mixed D(1)/D(2) dopaminergic agonist, apomorphine (APO; 0.25, 0.5 and 0.75 mg/kg sc) were tested on ASR and PPI of acoustic startle. Without APO, all groups of rats showed no difference in baseline ASR or PPI of acoustic startle. Following administration of APO (0.25, 0.5 and 0.75 mg/kg), ASR was significantly increased in OVX rats compared to intact rats and this was corrected with E(2) treatment. In all groups of animals, APO decreased PPI of acoustic startle. APO disrupted PPI to a lesser extent in OVX animals with or without E(2) treatment compared to intact rats. However, when group differences in APO-induced ASR were statistically controlled for, there were no longer any differences in APO disruption of PPI among the three treatment groups. These results indicate that long-term ovariectomy has persistent effects on the modulation of ASR, and these effects can be at least partly corrected with E(2) replacement therapy.

Comparisons of behavioral and neurochemical characteristics between WKY, WKHA, and Wistar rat strains.

WKHA rats constitute a recombinant inbred rat strain derived by phenotypic selection of the progeny of hybrid F2 crosses between SHR and WKY rats. WKHA are normotensive and show some features of hyperactivity and of hyper-reactivity to stress, but their utility as model of attention deficit/hyperactivity disorder (ADHD) has not yet been settled. To address these questions, we performed behavioral and neurochemical evaluations of WKHA, and compared them to both WKY and Wistar (WIS) rats. In locomotor activity tests, the respective scores for each strain were WKY

Fluoxetine-induced increases in open-field habituation in the olfactory bulbectomized rat depend on test aversiveness but not on anxiety.

Little is known regarding the functional processes underlying the treatment efficacy of antidepressant drugs. Given the close association between stress, anxiety and depression, distinguishing the common and disparate features of these processes may contribute to our current understanding. Using the olfactory bulbectomized (OBX) rat, an animal model sensitive to a variety of antidepressant drugs, this study examined the effects of chronic fluoxetine administration on open-field behavior under different conditions of stressfulness (luminance) and compared the fluoxetine effects to those evoked by the anxiolytic lorazepam. Sham-operated and OBX rats received 21 daily injections of fluoxetine (10 mg/kg), one or seven injections of lorazepam (0.1 and 0.5 mg/kg) or vehicle prior to testing in the open field or plus maze. Time series data were collected and fit with exponential regression models to estimate behavioral reactivity, habituation and residual rate of responding. Relative to sham controls, OBX rats displayed increased locomotor activity in the high luminance open field but showed decreased activity in the lower luminance open field. Time series analysis revealed that while sham animals showed increased habituation in the high compared to lower luminance open field, OBX rats did not significantly modify their responding between the two conditions. Chronic fluoxetine treatment invoked rectifying effects in OBX animals only in the high luminance open field by increasing the rate of habituation. Both acute and subchronic administration of lorazepam also reduced OBX hyperactivity but did so only by decreasing the residual rate of responding. As expected, lorazepam administration significantly increased the ratio of open-to-total arm activity in the elevated plus maze. These findings suggest that OBX responding in the open field may be maladaptive, reflecting an inability to modify behavior appropriately in certain environmental contexts. Chronic antidepressant treatment enhances habituation of OBX animals only under more stressful or aversive conditions and appears to do so in a manner temporally distinct from anxiolytic treatment.

Suckling-induced changes in responsivity to the hypoalgesic effect of morphine.

The hypoalgesic effect of morphine in lactating rats was assessed using the hot-plate test. At midlactation (days 12 and 18 postpartum) females nursing litters of 8 pups were less responsive to the hypoalgesic effect of morphine than ovariectomised or cycling females and females on days 6 or 24 of lactation. Subsequent studies showed that the hypoalgesic response to morphine was inhibited in lactating rats at a number of time points after drug administration and across a variety of doses. This effect was not dependent on milk delivery but was dependent on the hormonal state of the female since separation of dams and their litters for 96 h was sufficient to reinstate the response to morphine if it resulted in a reappearance of vaginal estrus.