Top2a promotes the development of social behavior via PRC2 and H3K27me3.

Little is understood about the embryonic development of sociality. We screened 1120 known drugs and found that embryonic inhibition of topoisomerase IIα (Top2a) resulted in lasting social deficits in zebrafish. In mice, prenatal Top2 inhibition caused defects in social interaction and communication, which are behaviors that relate to core symptoms of autism. Mutation of Top2a in zebrafish caused down-regulation of a set of genes highly enriched for genes associated with autism in humans. Both the Top2a-regulated and autism-associated gene sets have binding sites for polycomb repressive complex 2 (PRC2), a regulatory complex responsible for H3K27 trimethylation (H3K27me3). Moreover, both gene sets are highly enriched for H3K27me3. Inhibition of the PRC2 component Ezh2 rescued social deficits caused by Top2 inhibition. Therefore, Top2a is a key component of an evolutionarily conserved pathway that promotes the development of social behavior through PRC2 and H3K27me3.

Seroprevalence of Q fever in cattle, sheep and goats in the Volta region of Ghana.

Q fever is a zoonotic disease caused by Coxiella burnetii, a causative agent of abortion in livestock and febrile illness in humans. Outbreaks of human cases of Q fever have been reported in Australia and the Netherlands, which was linked to abortions in goat and sheep farms. In Ghana, information on Q fever in both livestock and humans is scanty. This study sought to determine the seroprevalence of Q fever in livestock in the Tongu area of the Volta region of Ghana. It was a cross sectional study with blood sampled from 204 cattle, 158 sheep and 100 goats. An indirect ELISA test was performed to detect Q fever antibodies in the serum of livestock. A total of 20 farms were sampled across the municipalities and an overall prevalence of Q fever was 21.6%. Specie-specific prevalence was 28.4% (45/158) for sheep, 21.7% (45/204) for cattle and 10% (10/100) for goats. Abortions were reported on all the farms sampled and most farmers lived in close proximity to the farms sampled. Q fever is prevalent in the North Tongu area and requires the attention of the veterinary and health authorities, using the One- Health approach in order to control its occurrence and save lives.

Dose-dependency of life span prolongation of F344/DuCrj rats injected with (-)deprenyl.

The effect of (-)deprenyl (D) on prolonging survival has previously been reported in different species of animals. In rats, three studies reported a positive effect, while one study reported a shortening of life spans. In the present study, we attempted to clarify past discrepancies in the results based on the speculation that there exists a certain effective dose range for this effect of the drug. F344/DuCrj rats of both sexes began to receive subcutaneous (s.c.) injections of D at the age of 18 months at a dose of 0.25 mg/kg/injection (inj.), 3 times a week. Control animals were given a vehicle (a saline solution). Average life spans of animals (days) were significantly increased in both male (895 +/- 109.7, n=30; 967.8 +/- 88.6, n=30, control vs. D treated, P<0.01, t-test) and female (924.7 +/- 132.2, n=38; 987.1 +/- 133.4, n=39, P<0.05) rats by 8.1% and 6.7%, respectively. We have previously reported that a dose of 0.5mg/kg/inj. (s.c.) significantly increased the life span of male F344 rats, while a dose of 1.0 mg/kg/inj. somewhat shortened the life span, although the difference was not statistically significant. The results of the present study coupled with our previous reports clearly indicate that a proper dose of D within a certain dose range can significantly increase the life span of animals of both sexes, but that a greater dose becomes less effective and may actually adversely affect the life span of rats. The presence of this effective dose range of D may explain discrepancies in the effect of D on life spans of animals previously reported.

Deletion of Pten in mouse brain causes seizures, ataxia and defects in soma size resembling Lhermitte-Duclos disease.

Initially identified in high-grade gliomas, mutations in the PTEN tumor-suppressor are also found in many sporadic cancers and a few related autosomal dominant hamartoma syndromes. PTEN is a 3'-specific phosphatidylinositol-3,4,5-trisphosphate (PI(3,4,5)P3) phosphatase and functions as a negative regulator of PI3K signaling. We generated a tissue-specific deletion of the mouse homolog Pten to address its role in brain function. Mice homozygous for this deletion (PtenloxP/loxP;Gfap-cre), developed seizures and ataxia by 9 wk and died by 29 wk. Histological analysis showed brain enlargement in PtenloxP/loxP;Gfap-cre mice as a consequence of primary granule-cell dysplasia in the cerebellum and dentate gyrus. Pten mutant cells showed a cell-autonomous increase in soma size and elevated phosphorylation of Akt. These data represent the first evidence for the role of Pten and Akt in cell size regulation in mammals and provide an animal model for a human phakomatosis condition, Lhermitte-Duclos disease (LDD).

Do antioxidant strategies work against aging and age-associated disorders? Propargylamines: a possible antioxidant strategy.

The free radical theory of aging was initially proposed by Harman half a century ago primarily to explain biological aging processes. Although administration of so-called antioxidant chemicals, which have been tested in the past for several decades, turned out to be mostly ineffective in prolonging the life spans of animals, the same theory of age-associated diseases appears to be increasingly supported in the last two decades. Despite these difficulties, the success in extending life span of 4 different animal species (mice, rats, hamsters, and dogs) with (-)deprenyl (including a study of our group) indicates that there might exist another type of antioxidant strategy in addition to a simple administration of antioxidant chemicals. (-)Deprenyl has also been shown to increase superoxide dismutase (SOD) and catalase (CAT) activities selectively in brain dopaminergic tissues. Interestingly, we have recently shown that another propargylamine, rasagiline not only increases antioxidant enzyme activities (CAT and SOD) in brain dopaminergic regions as (-)deprenyl does, but also increases CAT and SOD activities in extrabrain catecholaminergic systems such as the heart and kidneys as well. These recent observations coupled with previous observations on the life span of animals with (-)deprenyl suggest that pharmacological modulation of endogenous antioxidant enzyme activities could be one potential antioxidant strategy against aging and age-associated disorders. If the causal relationship between the two effects of (-)deprenyl exists as we hypothesized, we might be able to advance the elucidation of mechanism(s) of aging based on the free radical theory of aging.

A high dose of long term treatment with deprenyl loses its effect on antioxidant enzyme activities as well as on survivals of Fischer-344 rats.

The survival rate of male Fischer-344/Du rats treated chronically with high doses of deprenyl was investigated. Eighteen month old rats were treated with 1 mg/kg s.c. deprenyl 3 times per week for 13 months. At the age of 31 months, treated rats showed a greater mortality rate with three of 12 rats surviving, while in saline-treated control animals seven of 12 animals survived. No significant differences in superoxide dismutase (SOD) or catalase (CAT) activities in brain regions of control and treated animals were seen at 31 months of age. In contrast, when 27 month old rats were treated in the same manner for one month, significant increases in SOD (both Cu,Zn- and Mn-) and CAT activities were found in substantia nigra, striatum and cerebral cortex, but not in hippocampus. This effect was produced with a wide range of deprenyl doses (0.25-2 mg/kg, but not 4 mg/kg). Although a causal relationship between the two different effects of the drug, i.e. 1) increases in antioxidant enzyme activities and 2) the prolongation of survival of animals, has not been directly demonstrated, the loss of both effects with the high dose of the drug in the present experiment may be taken as circumstantial evidence for their causal relationship.

Plasticity in the maternal circuit: effects of experience and partum condition on brain astrocyte number in female rats.

Female rats that have received a maternal experience undergo enhanced c-fos expression in a number of brain sites when reexposed to pups. The present 2 studies examined changes in the expression of another brain protein, glial fibrillary acidic protein (GFAP), which is a major unit of the astrocytic cytoskeleton. In both experiments, primiparous and multiparous female rats were given varying amounts of postpartum contact with pups and overdosed after varying intervals, with no pups. Brains were prepared for GFAP immunohistochemical analysis. In both studies, Day 5 postpartum multiparous subjects given additional postpartum contact with pups, when compared with pup-exposed primiparous subjects, were found to have significantly higher numbers of GFAP positive cells in the medial preoptic area of the hypothalamus, an area critical for the expression of maternal behavior, but not in control sites. In Experiment 2, an opposite effect of parity was found in the medial amygdala and habenula.

Common properties for propargylamines of enhancing superoxide dismutase and catalase activities in the dopaminergic system in the rat: implications for the life prolonging effect of (-)deprenyl.

(-)Deprenyl has been reported to prolong the life span of different animal species. Further, the drug effectively increases antioxidant enzyme activities such as superoxide dismutase (SOD) and catalase (CAT) in brain dopaminergic regions. We have found that the effect of the drug on antioxidant enzyme activities is highly dose dependent, increasing with an increasing dose, however, a higher dose becomes less effective and an excessive dose becomes adversely effective. Most importantly, an optimal dose for the effect varies widely depending on animal species, strain, sex, age and duration of the treatment, which may at least partly explain discrepancies reported among different studies in the past. From the parallelism of the dose-effect relationship of the drug between life span extension and increasing endogenous antioxidant enzyme activity, we have suggested that the above two effects of (-)deprenyl may be causally related. This review summarizes our past series of studies and also reports our very recent observation that other propargylamines such as rasagiline and (R)-N-(2-heptyl)-N-methylpropagylamine (R-2HMP) also share the property of enhancing antioxidant enzyme activities. Further, our most recent study has found that these propargylamines increase antioxidant enzyme activities not only in brain dopaminergic regions but in extra-brain dopaminergic tissues such as the heart and kidneys. These observations are discussed in relation to the life prolonging effect of (-)deprenyl reported in the past.

Induction of SC1 mRNA encoding a brain extracellular matrix glycoprotein related to SPARC following lesioning of the adult rat forebrain.

SC1 is an extracellular matrix (ECM) glycoprotein related to SPARC which exhibits anti-adhesive properties. ECM molecules are thought to play important roles in influencing cell shape, proliferation and migration during neurogenesis. Following localized injury to the adult rat forebrain, a biphasic induction of SC1 mRNA was apparent, namely a rapid, transient induction at 1 day post-lesion in cortical neurons which border the lesion site followed by a more prolonged induction in astrocytes which are proximal to the wound site. A similar SC1 induction pattern was observed in the hippocampus in response to the injury. SPARC mRNA exhibits a divergent pattern of induction because it is induced in mature blood vessels close to the lesion and in blood vessels which develop following the trauma. Thus mRNAs encoding the related ECM glycoproteins SC1 and SPARC are induced in different cell populations in the adult forebrain during the neural response to localized injury.

Pharmacological modifications of endogenous antioxidant enzymes with special reference to the effects of deprenyl: a possible antioxidant strategy.

Limited information is available on the upregulation of endogenous antioxidant enzymes by means of administering various pharmaceuticals and/or chemicals. It has been reported that ursodeoxycholic acid (UDCA), a bile acid originally identified from black bear bile (a Chinese medicine, Yutan) increased glutathione S-transferase (GST) activities in mouse livers, resulting in a decrease in systemic lethal toxicity of orally challenged 1-2-dichloro-4-nitrobenzene (DCNB). Also, ursolic acid found in herbal medicines (e.g. leaves of loquat) was reported to increase catalase (CAT) activities in mouse liver. Interestingly, the chemical structures of these two compounds are surprisingly similar to each other, despite the difference in their original sources. These results suggest that in the future, more and more compounds will be found to have effects on increasing endogenous antioxidant enzyme activities. Deprenyl is a monoamine oxidase B inhibitor but also possesses many other different pharmacological activities. Among these various pharmacological effects of deprenyl, a possible causal relationship between two effects of deprenyl, namely the prolongation of the survival of animals and upregulation of antioxidant enzymes in selective brain regions, has been postulated by the authors. In at least four different animal species (rats, mice, hamsters and dogs), a significant prolongation of survival by chronic administration of the drug has been reported by different groups including that of the authors. This group has reported that repeated administration of the drug for 2-3 weeks can significantly increase activities of both types of superoxide dismutase (SODs) (Cu, Zn-, and Mn-SODs) as well as of CAT selectively in brain dopaminergic regions. Both effects are dose dependent but excessive dosages become less effective and even cause an adverse effect (i.e. a decrease in enzyme activities and shortening of life span). The parallelism of the dose-effect relationship between the two phenomena suggests that modification of SOD and CAT levels is one possible mechanism for deprenyl's ability to prolong the life span of animals.

Time course for kindling-induced changes in the hilar area of the dentate gyrus: reactive gliosis as a potential mechanism.

Recurrent seizure activity induced during kindling has been reported to cause an increase in the hilar area of the dentate gyrus of the hippocampus. To date, very little is known about the mechanism of this increase. This study investigated the time course for kindling-induced changes in the hilar area of the dentate gyrus at seven days, one month, and two months post-kindling. Hilar area of the dentate gyrus was significantly increased by approximately 46% at seven days and remained elevated at one month, but declined back to control levels by two months. Glial fibrillary acidic protein (GFAP) immunostaining was also evaluated at the same time points to determine whether kindling-induced changes in the hilar area of the dentate gyrus are related to kindling-induced glial cell changes. Increases in hilar GFAP immunostaining by approximately 57% were observed at seven days and at one month post-kindling, but not at two months post-kindling. These findings indicate that kindling-induced changes in the hilar area of the dentate gyrus and kindling-induced glial cell changes follow a similar time course, and that kindling-induced glial cell changes may mediate the observed changes in the hilar area of the dentate gyrus.

SPARC/osteonectin mRNA is induced in blood vessels following injury to the adult rat cerebral cortex.

Recently we described the pattern of expression of the anti-adhesive glycoprotein SPARC/osteonectin in the developing and adult brain. SPARC mRNA was present in developing blood vessels during neurogenesis, but was not detected in the mature vasculature. We have now examined the effect of a lesion to the adult rat cerebral cortex on the expression of SPARC by in situ hybridization. SPARC mRNA was increased in the zone proximal to the wound at 3 to 10 days after cortical brain injury. During this period, SPARC was induced in mature blood vessels close to the lesion site and in blood vessels which develop following injury. These results suggest a role for SPARC in the process of angiogenesis following injury to the adult cerebral cortex.

Assessing the effects of deprenyl on longevity and antioxidant defenses in different animal models.

Among many pharmaceuticals that have been tested for their effects on longevities of different animal rodents, deprenyl is unique in that its effects on longevity has been tested in at least four different animal species by independent research groups and that the effect has been postulated to be due to its effect of raising such antioxidant enzyme activities as superoxide dismutase (SOD) and catalase (CAT) in selective brain regions. Thus far, in all four species of animals examined (rats, mice, hamsters, and dogs), a positive effect was demonstrated, although the extent of its effect is quite variable. Our group has examined the effect on longevities in rats and mice and on antioxidant enzymes in rats, mice, and dogs. Although in rats of both sexes, we have obtained positive effects on longevity, two studies with different doses in mice did not reveal a significantly positive effect. We have observed, however, significantly positive effects on SOD (in Cu, Zn-, and Mn-) as well as CAT (but not glutathione peroxidase) activities in the brain dopaminergic system such as in the S. nigra and striatum (but not in hippocampus) in all rats, mice, and dogs, although the effects were quite variable, depending on the doses used. In mice, however, a long-term administration (3x/w, 3 months) caused a remarkable decrease in the magnitude of activity as well as a narrowing of the effective dose range, which may explain a relatively weak effect of the drug on mouse longevity. Further, a recent study on aging beagle dogs by Ruehl et al. showed a remarkable effect on longevity, which agrees with our SOD study in dogs. Although deprenyl has been claimed to have several other effects, such as a radical scavenging effect and a neuroprotective effect, past reports on its effects on longevities and antioxidant defenses are compatible with the notion that the drug prolongs the life span of animals by reducing the oxidative damage to the brain dopaminergic system during aging. Further, our studies on F-344 rats as well as a dog study by Ruehl et al. suggest that the drug may at least partially prolong the life span of animals by enhancing immune system function and preventing tumor development in animals.

Chloroquine administration in mice increases beta-amyloid immunoreactivity and attenuates kainate-induced blood-brain barrier dysfunction.

The anti-malarial drug chloroquine (CHL) has been reported to cause the accumulation of beta-amyloid peptide containing fragments (fA beta) of the amyloid precursor protein within lysosomes in vitro. However, the significance of this finding with regards to the development of Alzheimer's disease (AD) pathology in vivo is not known. Hence, we investigated the effects of chronic CHL administration in the mouse. Systemically administered CHL caused an astrocytic response and an increase in intracellular A beta immunoreactivity throughout the brain, but no plaque-like pathology. Pharmacological challenge with the excitotoxin kainic acid (KA) revealed a mild proconvulsant effect of CHL pretreatment (P < 0.06). Interestingly, CHL protected the blood-brain barrier from characteristic KA-induced dysfunction. Given the hypothesized involvement of both excitotoxic processes and the vascular system in AD, the observed interactions may assist in elucidating the pathogenesis of AD.

Long-term treatment of male F344 rats with deprenyl: assessment of effects on longevity, behavior, and brain function.

L-Deprenyl (selegiline) was chronically administered to male Fischer 344 rats via their drinking water beginning at 54 weeks of age (estimated daily dose: 0.5 mg/kg/day). Beginning at 84 weeks of age, the rats were behaviorally evaluated using a sensorimotor battery, a motor-learning task, and the Morris water maze. At 118 weeks of age, cerebellar noradrenergic function was evaluated in the surviving rats using in vivo electrochemistry. The rats were then sacrificed to measure brain monoamine oxidase activity and perform quantitative autoradiography to evaluate the effect of chronic deprenyl treatment on beta-adrenergic receptors in the cerebellum, alpha 2-adrenergic receptors several brain regions, and D1 and D2 dopamine receptors in the striatum. Deprenyl treatment reduced brain monoamine oxidase B activity by 85%, but had no effect on brain monoamine oxidase A. A clear effect of chronic deprenyl treatment upon longevity was not observed. Several measures of CNS function were altered in the deprenyl-treated animals: 1) spatial learning in the Morris water maze was improved; 2) electrochemical signals recorded following local application of NE were reduced, and the responsiveness to the reuptake blocker nomifensine was enhanced, in the cerebellum; 3) beta-adrenergic receptor binding affinity was increased in the cerebellum; 4) alpha 2-adrenergic receptor density was increased in the inferior colliculus; and 5) striatal D1 dopamine receptor density was reduced but binding affinity was enhanced. In contrast, chronic deprenyl treatment did not cause changes in: 1) sensorimotor function, as evaluated by balance beam, inclined screen, or wire hang tasks; 2) motor learning; 3) alpha 2-adrenergic receptor density in any region examined except for the inferior colliculus, or binding affinity in any region examined; or 4) striatal D2 dopamine receptor number or affinity. Thus, long-term oral administration of deprenyl extended the functional life span of rats with respect to cognitive, but not motor, performance.

Astrocytes in kindling: relevance to epileptogenesis.

Astrogliosis is a prominent feature of epileptic foci, and may play a causal role in the development of seizures and the persistance of seizure disorders. We have studied morphological changes in astrocytes with respect to the evolution of seizures using the kindling model of epilepsy. Kindling-induced seizures result in a prominent hypertrophy of astrocytes that is accompanied by a reorganization of astrocytic cytoskeleton. The change in the morphology of astrocytes appears to be seizure-intensity dependent, occurs early in the kindling process, and persists for weeks following the last seizure. In addition to hypertrophy, we have observed an increase in proliferation of astrocytes in hippocampus, amygdala and piriform cortex, but no change in the expression of connexin-43 following kindling. Significantly, induction of a localized astrocyte hypertrophy prior to initiation of kindling does not result in seizures and does not facilitate kindling. Altogether these data suggest that 'gliosis' is an adaptive response to seizures.

SC1, a brain extracellular matrix glycoprotein related to SPARC and follistatin, is expressed by rat cerebellar astrocytes following injury and during development.

In the nervous system, extracellular matrix components are believed to influence cell shape, proliferation and migration during development and following injury. SC1 is a secreted glycoprotein expressed during neural development and in the adult brain. The molecule shows partial sequence homology to the anti-adhesive extracellular matrix molecule SPARC/osteonectin and to follistatin. We have made a surgical lesion in the adult rat cerebellum and examined changes in SC1 expression at 1 to 14 days after injury. Dual in situ hybridization/immunohistochemistry demonstrated that SC1 mRNA was induced in astrocytes surrounding the wound, reaching maximal levels at 10 days post-lesion. Immunohistochemistry revealed changes in the deposition of SC1 protein in radial fibres of Bergmann glia. SC1 protein was also detected at the border of the lesion, suggesting an association with the glial scar. Double immunohistochemistry with the astrocytic marker GFAP demonstrated that astrocytes also express SC1 during postnatal development.

Short intertrial intervals impair water maze performance in old Fischer 344 rats.

Male and female Fischer 344 rats (N = 55) aged approximately 18, 21, and 24 months were tested for spatial learning in the water maze with intertrial intervals of 1-4 min (Massed) or 23-33 min (Spaced). Animals tested in the Massed condition showed an age-related impairment on trials to criterion; rats aged 24 months performed more poorly than younger subjects. Spaced animals did not differ at any age nor did they differ from 18- or 21-month-old Massed subjects. The youngest rats in both groups were comparable to animals aged 7-8 months tested under Massed conditions. Tests on swim distance, swim speed, and escape latency produced similar results. Our data suggest that acquisition deficits in 24-month-old rats tested with long intertrial intervals are due at least in part to increased susceptibility to fatigue and/or thermal stress. Caution should be used, therefore, when interpreting age-related impairments in water maze performance.

Leupeptin causes an accumulation of lipofuscin-like substances and other signs of aging in kidneys of young rats: further evidence for the protease inhibitor model of aging.

The "protease inhibitor model of aging" has been proposed on the basis of observations that young rat brains, livers, and retinas exposed to a protease inhibitor, leupeptin, accumulate lipofuscin-like substances (LLS) that are similar to age pigment, and also display a variety of other manifestations of aging. In order to validate this hypothesis in more general terms, the present study reports attempts to induce age-like changes in kidney cells of young rats. Male F-344 rats (4-5 weeks of age) were continuously infused intraperitoneally (i.p.) with various doses of leupeptin (1-50 mg/100 g/day) for 2 weeks. Control animals received saline solution and normal aged rats (27-30 months-old) received no treatment. Animals were sacrificed and subjected to histological examination. In kidneys of leupeptin-, but not saline-treated rats, generally round-shaped PAS-positive particles were clearly observed, which were predominantly distributed in proximal convoluted tubules, and which resembled particles in normal aged kidneys. With increasing drug doses, particles tended to become bigger and more numerous. The dominant accumulation of LLS in cells of the proximal convoluted tubules had a fine morphologic configuration that resembled age pigments in old rats. Also, there was a concomitant thickening of the basement membrane that was present in leupeptin-treated and aged kidneys, but not in controls. The results, therefore, support the protease inhibitor model of aging and provide an experimental tool for probing the cellular mechanisms of aging.

Upregulation of antioxidant enzyme activities by deprenyl. Implications for life span extension.

In order to elucidate the exact role of antioxidant enzyme activities such as superoxide dismutase (SOD) in the aging process of animals, we compared various enzyme activities in different brain regions and in the liver of young (6-8 mo) and old (28-30 mo) Fischer-344 (F-344) rats. While Mn-SOD activities were elevated 3-5-fold in specific brain regions such as hippocampus, striatum and substantia nigra in brains of old male rats compared with the young, in females both forms of SOD (Cu, Zn- and Mn-) enzyme activities remained essentially unchanged with aging. Continued subcutaneous infusion of deprenyl for 3 weeks caused a 2-3-fold increase in activities of both Cu Zn- and Mn-SOD and a 50-60% increase in CAT activities in striatum and substantia nigra but not in hippocampus, cerebellum or the liver. Further, long-term treatment of old male rats with deprenyl caused a significant increase in the remaining life expectancy from 24 months of age by 34%. In conclusion, activities of antioxidant enzymes in these regions examined do not show any uniform age-associated change, suggesting that changes in these enzyme activities do not have any specific role in the life span of rodents in general terms. In contrast, the results of our deprenyl study suggests the possibility that the protection of catecholaminergic neurons by an upregulation of SOD and CAT activities plays a significant role in the life span of animals.