Aberrant Somatosensory Processing and Connectivity in Mice Lacking Engrailed-2.

Overreactivity and defensive behaviors in response to tactile stimuli are common symptoms in autism spectrum disorder (ASD) patients. Similarly, somatosensory hypersensitivity has also been described in mice lacking ASD-associated genes such as Fmr1 (fragile X mental retardation protein 1). Fmr1 knock-out mice also show reduced functional connectivity between sensory cortical areas, which may represent an endogenous biomarker for their hypersensitivity. Here, we measured whole-brain functional connectivity in Engrailed-2 knock-out (En2-/-) adult mice, which show a lower expression of Fmr1 and anatomical defects common to Fmr1 knock-outs. MRI-based resting-state functional connectivity in adult En2-/- mice revealed significantly reduced synchronization in somatosensory-auditory/associative cortices and dorsal thalamus, suggesting the presence of aberrant somatosensory processing in these mutants. Accordingly, when tested in the whisker nuisance test, En2-/- but not WT mice of both sexes showed fear behavior in response to repeated whisker stimulation. En2-/- mice undergoing this test exhibited decreased c-Fos-positive neurons (a marker of neuronal activity) in layer IV of the primary somatosensory cortex and increased immunoreactive cells in the basolateral amygdala compared with WT littermates. Conversely, when tested in a sensory maze, En2-/- and WT mice spent a comparable time in whisker-guided exploration, indicating that whisker-mediated behaviors are otherwise preserved in En2 mutants. Therefore, fearful responses to somatosensory stimuli in En2-/- mice are accompanied by reduced basal connectivity of sensory regions, reduced activation of somatosensory cortex, and increased activation of the basolateral amygdala, suggesting that impaired somatosensory processing is a common feature in mice lacking ASD-related genes.SIGNIFICANCE STATEMENT Overreactivity to tactile stimuli is a common symptom in autism spectrum disorder (ASD) patients. Recent studies performed in mice bearing ASD-related mutations confirmed these findings. Here, we evaluated the behavioral response to whisker stimulation in mice lacking the ASD-related gene Engrailed-2 (En2-/- mice). Compared with WT controls, En2-/- mice showed reduced functional connectivity in the somatosensory cortex, which was paralleled by fear behavior, reduced activation of somatosensory cortex, and increased activation of the basolateral amygdala in response to repeated whisker stimulation. These results suggest that impaired somatosensory signal processing is a common feature in mice harboring ASD-related mutations.

Hypertension Impairs Cerebral Blood Flow in a Mouse Model for Alzheimer's Disease.

AIMS: Hypertension, a risk factor for Alzheimer's disease (AD), is a treatable condition, which offers possibilities for prevention of AD. Elevated angiotensin II (AngII) is an important cause of essential hypertension. AngII has deleterious effects on endothelial function and cerebral blood flow (CBF), which may contribute to AD. AngII blocking agents can thus provide potential candidates to reduce AD risk factors in hypertensive patients. METHODS: We studied the effect of 2 months induced hypertension (AngII-infusion via osmotic micropumps) on systolic blood pressure (SBP) and CBF in 10 months-old wild-type (WT) C57bl/6j and AßPPswe/PS1ΔE9 (AßPP/PS1) mice, and treatment with two different antihypertensives, 1) eprosartan mesylate (EM, 0.35mg/kg) or 2) hydrochlorotiazide (HCT, 7.5mg/kg), after 1 month of induced-hypertension. SBP was monitored twice each month via tail cuff plethysmography. CBF was measured with MR by flow-sensitive alternating inversion recovery. RESULTS: Chronic AngII-infusion induced an increase in SBP in both AßPP/PS1 and WT mice accompanied by a decrease in hippocampal and thalamic CBF only in the AßPP/PS1 mice. An additional difference between the AßPP/PS1 mice and WT mice was that SBP was much higher in AßPP/PS1 mice in both hypertensive and normotensive conditions. Moreover, both antihypertensives were less effective in reducing AngII-induced hypertension to normal levels in AßPP/PS1 mice, while being effective in WT mice. CONCLUSIONS: It can be concluded that AngII-induced elevated SBP results in impaired CBF and a decreased response to blood pressure lowering treatment in a transgenic model of AD. Our findings suggest a relation between midlife hypertension and decreased CBF in an AD mouse model, similar to the relation which has been found in AD patients. This translational mouse model could be used to investigate possible prevention and treatment strategies for AD.

Effect of perinatally supplemented flavonoids on brain structure, circulation, cognition, and metabolism in C57BL/6J mice.

Evidence suggests that flavanol consumption can beneficially affect cognition in adults, but little is known about the effect of flavanol intake early in life. The present study aims to assess the effect of dietary flavanol intake during the gestational and postnatal period on brain structure, cerebral blood flow (CBF), cognition, and brain metabolism in C57BL/6J mice. Female wild-type C57BL/6J mice were randomly assigned to either a flavanol supplemented diet or a control diet at gestational day 0. Male offspring remained on the corresponding diets throughout life and performed cognitive and behavioral tests during puberty and adulthood assessing locomotion and exploration (Phenotyper and open field), sensorimotor integration (Rotarod and prepulse inhibition), and spatial learning and memory (Morris water maze, MWM). Magnetic resonance spectroscopy and imaging at 11.7T measured brain metabolism, CBF, and white and gray matter integrity in adult mice. Biochemical and immunohistochemical analyses evaluated inflammation, synaptic plasticity, neurogenesis, and vascular density. Cognitive and behavioral tests demonstrated increased locomotion in Phenotypers during puberty after flavanol supplementation (p = 0.041) but not in adulthood. Rotarod and prepulse inhibition demonstrated no differences in sensorimotor integration. Flavanols altered spatial learning in the MWM in adulthood (p = 0.039), while spatial memory remained unaffected. Additionally, flavanols increased diffusion coherence in the visual cortex (p = 0.014) and possibly the corpus callosum (p = 0.066) in adulthood. Mean diffusion remained unaffected, a finding that corresponds with our immunohistochemical data showing no effect on neurogenesis, synaptic plasticity, and vascular density. However, flavanols decreased CBF in the cortex (p = 0.001) and thalamus (p = 0.009) in adulthood. Brain metabolite levels and neuroinflammation remained unaffected by flavanols. These data suggest that dietary flavanols results in subtle alterations in brain structure, locomotor activity and spatial learning. Comparison of these data to published findings in aging or neurodegeneration suggests that benefits of dietary flavanols may increase with advancing age and in disease.

Impact of dietary n-3 polyunsaturated fatty acids on cognition, motor skills and hippocampal neurogenesis in developing C57BL/6J mice.

Maternal intake of omega-3 polyunsaturated fatty acids (n-3 PUFA) is critical during perinatal development of the brain. Docosahexaenoic acid (DHA) is the most abundant n-3 PUFA in the brain and influences neuronal membrane function and neuroprotection. The present study aims to assess the effect of dietary n-3 PUFA availability during the gestational and postnatal period on cognition, brain metabolism and neurohistology in C57BL/6J mice. Female wild-type C57BL/6J mice at day 0 of gestation were randomly assigned to either an n-3 PUFA deficient diet (0.05% of total fatty acids) or an n-3 PUFA adequate diet (3.83% of total fatty acids) containing preformed DHA and its precursor α-linolenic acid. Male offspring remained on diet and performed cognitive tests during puberty and adulthood. In adulthood, animals underwent (31)P magnetic resonance spectroscopy to assess brain energy metabolites. Thereafter, biochemical and immunohistochemical analyses were performed assessing inflammation, neurogenesis and synaptic plasticity. Compared to the n-3 PUFA deficient group, pubertal n-3 PUFA adequate fed mice demonstrated increased motor coordination. Adult n-3 PUFA adequate fed mice exhibited increased exploratory behavior, sensorimotor integration and spatial memory, while neurogenesis in the hippocampus was decreased. Selected brain regions of n-3 PUFA adequate fed mice contained significantly lower levels of arachidonic acid and higher levels of DHA and dihomo-γ-linolenic acid. Our data suggest that dietary n-3 PUFA can modify neural maturation and enhance brain functioning in healthy C57BL/6J mice. This indicates that availability of n-3 PUFA in infant diet during early development may have a significant impact on brain development.

Impact of a multi-nutrient diet on cognition, brain metabolism, hemodynamics, and plasticity in apoE4 carrier and apoE knockout mice.

Lipid metabolism and genetic background together strongly influence the development of both cardiovascular and neurodegenerative diseases like Alzheimer's disease (AD). A non-pharmacological way to prevent the genotype-induced occurrence of these pathologies is given by dietary behavior. In the present study, we tested the effects of long-term consumption of a specific multi-nutrient diet in two models for atherosclerosis and vascular risk factors in AD: the apolipoprotein ε4 (apoE4) and the apoE knockout (apoE ko) mice. This specific multi-nutrient diet was developed to support neuronal membrane synthesis and was expected to contribute to the maintenance of vascular health. At 12 months of age, both genotypes showed behavioral changes compared to control mice and we found increased neurogenesis in apoE ko mice. The specific multi-nutrient diet decreased anxiety-related behavior in the open field, influenced sterol composition in serum and brain tissue, and increased the concentration of omega-3 fatty acids in the brain. Furthermore, we found that wild-type and apoE ko mice fed with this multi-nutrient diet showed locally increased cerebral blood volume and decreased hippocampal glutamate levels. Taken together, these data suggest that a specific dietary intervention has beneficial effects on early pathological consequences of hypercholesterolemia and vascular risk factors for AD.

Multinutrient diets improve cerebral perfusion and neuroprotection in a murine model of Alzheimer's disease.

Nutritional intervention may retard the development of Alzheimer's disease (AD). In this study we tested the effects of 2 multi-nutrient diets in an AD mouse model (APPswe/PS1dE9). One diet contained membrane precursors such as omega-3 fatty acids and uridine monophosphate (DEU), whereas another diet contained cofactors for membrane synthesis as well (Fortasyn); the diets were developed to enhance synaptic membranes synthesis, and contain components that may improve vascular health. We measured cerebral blood flow (CBF) and water diffusivity with ultra-high-field magnetic resonance imaging, as alterations in these parameters correlate with clinical symptoms of the disease. APPswe/PS1dE9 mice on control diet showed decreased CBF and changes in brain water diffusion, in accordance with findings of hypoperfusion, axonal disconnection and neuronal loss in patients with AD. Both multinutrient diets were able to increase cortical CBF in APPswe/PS1dE9 mice and Fortasyn reduced water diffusivity, particularly in the dentate gyrus and in cortical regions. We suggest that a specific diet intervention has the potential to slow AD progression, by simultaneously improving cerebrovascular health and enhancing neuroprotective mechanisms.

Effects of specific multi-nutrient enriched diets on cerebral metabolism, cognition and neuropathology in AßPPswe-PS1dE9 mice.

Recent studies have focused on the use of multi-nutrient dietary interventions in search of alternatives for the treatment and prevention of Alzheimer's disease (AD). In this study we investigated to which extent long-term consumption of two specific multi-nutrient diets can modulate AD-related etiopathogenic mechanisms and behavior in 11-12-month-old AßPPswe-PS1dE9 mice. Starting from 2 months of age, male AßPP-PS1 mice and wild-type littermates were fed either a control diet, the DHA+EPA+UMP (DEU) diet enriched with uridine monophosphate (UMP) and the omega-3 fatty acids docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), or the Fortasyn® Connect (FC) diet enriched with the DEU diet plus phospholipids, choline, folic acid, vitamins and antioxidants. We performed behavioral testing, proton magnetic resonance spectroscopy, immunohistochemistry, biochemical analyses and quantitative real-time PCR to gain a better understanding of the potential mechanisms by which these multi-nutrient diets exert protective properties against AD. Our results show that both diets were equally effective in changing brain fatty acid and cholesterol profiles. However, the diets differentially affected AD-related pathologies and behavioral measures, suggesting that the effectiveness of specific nutrients may depend on the dietary context in which they are provided. The FC diet was more effective than the DEU diet in counteracting neurodegenerative aspects of AD and enhancing processes involved in neuronal maintenance and repair. Both diets elevated interleukin-1ß mRNA levels in AßPP-PS1 and wild-type mice. The FC diet additionally restored neurogenesis in AßPP-PS1 mice, decreased hippocampal levels of unbound choline-containing compounds in wild-type and AßPP-PS1 animals, suggesting diminished membrane turnover, and decreased anxiety-related behavior in the open field behavior. In conclusion, the current data indicate that specific multi-nutrient diets can influence AD-related etiopathogenic processes. Intervention with the FC diet might be of interest for several other neurodegenerative and neurological disorders.

Electrophysiologic efficacy of irrigated bipolar radiofrequency in the clinical setting.

OBJECTIVE: Clinical success of atrial fibrillation ablation depends on persistent transmurality of the lesions. Although bipolar radiofrequency grants acute pulmonary vein isolation, the fate of such ablations in the clinical setting is unknown. We assessed postoperative pulmonary vein isolation up to 3 weeks after open chest bipolar radiofrequency ablation. METHODS: Thirteen consecutive patients with mitral valve disease (mean age, 60 +/- 10 years) and atrial fibrillation undergoing concomitant ablation with the BP2 bipolar device (Medtronic, Inc, Minneapolis, Minn) were enrolled for electrophysiologic assessment. During surgery, pairs of additional temporary wires were positioned on the right pulmonary veins and on the roof of the left atrium before bipolar ablation. Entrance block (abatement or disconnection of electrogram potentials) and exit block (no entrainment during pulmonary vein pacing) of the right pulmonary veins and of the free left atrium were assessed before and after ablation. After right pulmonary vein isolation was obtained, one additional encircling line was added. Electrophysiologic assessment was repeated before discharge and at 3 weeks. RESULTS: Baseline right pulmonary vein pacing threshold was 2.9 +/- 1.6 mA. After 3 +/- 1 encircling ablations, bidirectional block was attained in all pulmonary veins. At pre-discharge electrophysiologic study, complete isolation persisted in all cases. At 3 weeks, conduction block persisted in 11 (85%) of 13 patients. All patients were discharged in sinus rhythm. At follow-up (19 +/- 7 months), 12 (92%) of 13 patients were still free from atrial fibrillation. CONCLUSIONS: Irrigated bipolar radiofrequency ablation provides acute transmurality after multiple ablations. However, total recovery of conduction occurred in 15% of the patients after 3 weeks. Repeated multiple ablations, possibly complemented by block validation, are suggested to help achieve durable transmurality with such technology.