Faculty Bibliography

Vascular endothelial growth factor (VEGF) is critical to angiogenesis and vascular permeability. It is also important in the endocrine system, in which VEGF mediates the vascular effects of estrogens in target tissues such as the uterus, a response attributed to an estrogen response element on the VEGF gene. Here we asked whether 17beta-estradiol increases VEGF levels in the brain. We focused on the hippocampus, in which 17beta-estradiol and VEGF both have important actions, and used immunocytochemistry to evaluate VEGF protein. VEGF immunoreactivity was compared in adult female rats sampled during the estrous cycle when serum levels of 17beta-estradiol peak (proestrous morning) as well as when they are low (metestrous morning). In addition, adult rats were ovariectomized and compared after treatment with 17beta-estradiol or vehicle. The results demonstrated that VEGF immunoreactivity was increased when serum levels of 17beta-estradiol were elevated. Confocal microscopy showed that VEGF immunofluorescence was predominantly nonneuronal, often associated with astrocytes. Glial VEGF labeling was primarily punctate rather than diffuse and labile because glial VEGF immunoreactivity was greatly reduced if tissue sections were left in an aqueous medium overnight. We conclude that VEGF protein in normal female hippocampus is primarily nonneuronal rather than neuronal and suggest that glial VEGF immunoreactivity has been underestimated by past studies with other methods because there is a labile extracellular pool. We suggest that estrogens may exert actions on female hippocampal structure and function by increasing hippocampal VEGF

Aquaporin-4 (AQP4) is the major water channel in the CNS and is primarily expressed in astrocytes. Little is known about the potential for AQP4 to influence synaptic plasticity, although many studies have shown that it regulates the response of the CNS to injury. Therefore, we evaluated long-term potentiation (LTP) and long-term depression (LTD) in AQP4 knock-out (KO) and wild-type mice. KO mice exhibited a selective defect in LTP and LTD without a change in basal transmission or short-term plasticity. Interestingly, the impairment in LTP in KO mice was specific for the type of LTP that depends on the neurotrophin BDNF, which is induced by stimulation at theta rhythm [theta-burst stimulation (TBS)-LTP], but there was no impairment in a form of LTP that is BDNF independent, induced by high-frequency stimulation. LTD was also impaired in KO mice, which was rescued by a scavenger of BDNF or blockade of Trk receptors. TrkB receptors, which mediate effects of BDNF on TBS-LTP, were not altered in KO mice, but p75NTR, the receptor that binds all neurotrophins and has been implicated in some types of LTD, was decreased. The KO mice also exhibited a cognitive defect, which suggests a new role for AQP4 and astrocytes in normal cognitive function. This defect was evident using a test for location-specific object memory but not Morris water maze or contextual fear conditioning. The results suggest that AQP4 channels in astrocytes play an unanticipated role in neurotrophin-dependent plasticity and influence behavior

Granule cell (GC) neurogenesis in the dentate gyrus (DG) does not always proceed normally. After severe seizures (e.g., status epilepticus [SE]) and some other conditions, newborn GCs appear in the hilus. Hilar ectopic GCs (EGCs) can potentially provide insight into the effects of abnormal location and seizures on GC development. Additionally, hilar EGCs that develop after SE may contribute to epileptogenesis and cognitive impairments that follow SE. Thus, it is critical to understand how EGCs differ from normal GCs. Relatively little morphometric information is available on EGCs, especially those restricted to the hilus. This study quantitatively analyzed the structural morphology of hilar EGCs from adult male rats several months after pilocarpine-induced SE, when they are considered to have chronic epilepsy. Hilar EGCs were physiologically identified in slices, intracellularly labeled, processed for light microscopic reconstruction, and compared to GC layer GCs, from both the same post-SE tissue and the NeuroMorpho database (normal GCs). Consistently, hilar EGC and GC layer GCs had similar dendritic lengths and field sizes, and identifiable apical dendrites. However, hilar EGC dendrites were topologically more complex, with more branch points and tortuous dendritic paths. Three-dimensional analysis revealed that, remarkably, hilar EGC dendrites often extended along the longitudinal DG axis, suggesting increased capacity for septotemporal integration. Axonal reconstruction demonstrated that hilar EGCs contributed to mossy fiber sprouting. This combination of preserved and aberrant morphological features, potentially supporting convergent afferent input to EGCs and broad, divergent efferent output, could help explain why the hilar EGC population could impair DG function

The Autism Tissue Program (ATP), a science program of Autism Speaks, provides researchers with access to well-characterized postmortem brain tissues. Researchers access these tissues through a peer-reviewed, project-based approval process, and obtain related clinical information from a secure, online informatics portal. However, few of these samples have DNA banked from other sources (such as a blood sample from the same individual), hindering genotype-phenotype correlation and interpretation of gene expression data derived from the banked brain tissue. Here, we describe an initiative to extract DNA from Brodmann Area 19, and genotype these samples using both the Affymetrix Genome-Wide Human SNP Array 6.0 and the Illumina Human1M-Duo DNA Analysis BeadChip genome-wide microarray technologies. We additionally verify reported gender, and infer ethnic background from the single nucleotide polymorphism data. We have also used a rigorous, multiple algorithm approach to identify genomic copy number variation (CNV) from these array data. Following an initial proof of principle study using two samples, 52 experimental samples, consisting of 27 subjects with confirmed or suspected autism and related disorders, 5 subjects with cytogenetically visible duplications of 15q, 2 with epilepsy and 18 age-matched normal controls were processed, yielding high-quality genotype data in all cases. The genotype and CNV data are provided via the ATP informatics portal as a resource for the autism research community

The neurofilament light subunit (NF-L) binds to myosin Va (Myo Va) in neurons but the sites of interaction and functional significance are not clear. We show by deletion analysis that motor domain of Myo Va binds to the NF-L rod domain that forms the NF backbone. Loss of NF-L and Myo Va binding from axons significantly reduces the axonal content of ER, and redistributes ER to the periphery of axon. Our data are consistent with a novel function for NFs as a scaffold in axons for maintaining the content and proper distribution of vesicular organelles, mediated in part by Myo Va. Based on observations that the Myo Va motor domain binds to intermediate filament (IF) proteins of several classes, Myo Va interactions with IFs may serve similar roles in organizing organelle topography in different cell types

The median eminence of the hypothalamus is an important conduit by which neurosecretory hormones from hypothalamic nuclei are delivered to the pars nervosa (neural lobe) of the pituitary en route tothe bloodstream. Dilutional hyponatremia was produced in adult ratsto determine the effect on the morphology of the median eminence of the hypothalamus. Hyponatremia was caused by reducing electrolyte and organic osmolyte reserves to block the excretion of water through delivery of the nephrotoxin mercuric chloride (HgCl2). Histological examination of the brain 1 day after a hyponatremic insult revealed vacuolation within the median eminence of the hypothalamus. No other lesions were found in other parts of the brain after hyponatremia. The hyponatremic lesion consisted of a band of closely packed vacuoles that crossed the floor of the third ventricle. Vacuoles associated with hyponatremia were predominantly in the subependymal, fiber, reticular, and palisade layers of the median eminence. Vacuolation was not observed in the tanycyte layer of the median eminence. This study indicates that the median eminence is a potentially vulnerable site in human hyponatremic conditions that should be evaluated further in relevant animal models

Neurodegeneration occurs in acute pathological conditions such as stroke, ischemia, and head trauma and in chronic disorders such as Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. While the cause of neuronal death is different and not always known in these varied conditions, hindrance of cell death would be beneficial in the prevention of, slowing of, or halting disease progression. Enhanced cystatin C (CysC) expression in these conditions caused a debate as to whether CysC up-regulation facilitates neurodegeneration or it is an endogenous neuroprotective attempt to prevent the progression of the pathology. However, recent in vitro and in vivo data have demonstrated that CysC plays protective roles via pathways that are dependent on inhibition of cysteine proteases, such as cathepsin B, or by induction of autophagy, induction of proliferation, and inhibition of amyloid-beta aggregation. Here we review the data demonstrating the protective roles of CysC under conditions of neuronal challenge and the protective pathways induced under various conditions. These data suggest that CysC is a therapeutic candidate that can potentially prevent brain damage and neurodegeneration.

Autophagy, a major degradative pathway for proteins and organelles, is essential for survival of mature neurons. Extensive autophagic-lysosomal pathology in Alzheimer's disease brain contributes to Alzheimer's disease pathogenesis, although the underlying mechanisms are not well understood. Here, we identified and characterized marked intraneuronal amyloid-beta peptide/amyloid and lysosomal system pathology in the Alzheimer's disease mouse model TgCRND8 similar to that previously described in Alzheimer's disease brains. We further establish that the basis for these pathologies involves defective proteolytic clearance of neuronal autophagic substrates including amyloid-beta peptide. To establish the pathogenic significance of these abnormalities, we enhanced lysosomal cathepsin activities and rates of autophagic protein turnover in TgCRND8 mice by genetically deleting cystatin B, an endogenous inhibitor of lysosomal cysteine proteases. Cystatin B deletion rescued autophagic-lysosomal pathology, reduced abnormal accumulations of amyloid-beta peptide, ubiquitinated proteins and other autophagic substrates within autolysosomes/lysosomes and reduced intraneuronal amyloid-beta peptide. The amelioration of lysosomal function in TgCRND8 markedly decreased extracellular amyloid deposition and total brain amyloid-beta peptide 40 and 42 levels, and prevented the development of deficits of learning and memory in fear conditioning and olfactory habituation tests. Our findings support the pathogenic significance of autophagic-lysosomal dysfunction in Alzheimer's disease and indicate the potential value of restoring normal autophagy as an innovative therapeutic strategy for Alzheimer's disease

Purpose: recent studies have found that KRAS mutations predict resistance to monoclonal antibodies targeting the epidermal growth factor receptor in metastatic colorectal cancer (mCRC). A polymorphism in a let-7 microRNA complementary site (lcs6) in the KRAS 3' untranslated region (UTR) is associated with an increased cancer risk in non-small-cell lung cancer and reduced overall survival (OS) in oral cancers. We tested the hypothesis whether this polymorphism may be associated with clinical outcome in KRAS wild-type (KRASwt) mCRC patients treated with cetuximab monotherapy. PATIENTS AND METHODS: the presence of KRAS let-7 lcs6 polymorphism was evaluated in 130 mCRC patients who were enrolled in a phase II study of cetuximab monotherapy (IMCL-0144). Genomic DNA was extracted from dissected formalin-fixed paraffin-embedded tumor tissue, KRAS mutation status and polymorphism were assessed using direct sequencing and PCR restriction fragment length polymorphism technique. RESULTS: KRAS let-7 lcs6 polymorphism was found to be related to object response rate (ORR) in mCRC patients whose tumors had KRASwt. The 12 KRASwt patients harboring at least a variant G allele (TG or GG) had a 42% ORR compared with a 9% ORR in 55 KRASwt patients with let-7 lcs6 TT genotype (P = 0.02, Fisher's exact test). KRASwt patients with TG/GG genotypes had trend of longer median progression-free survival (3.9 versus 1.3 months) and OS (10.7 versus 6.4 months) compared to those with TT genotypes. CONCLUSIONS: these results are the first to indicate that the KRAS 3'UTR polymorphism may predict for cetuximab responsiveness in KRASwt mCRC patients, which warrants validation in other clinical trials