Inhibition of neocortical beta-amyloid (Abeta) accumulation may be essential in an effective therapeutic intervention for Alzheimer's disease (AD). Cu and Zn are enriched in Abeta deposits in AD, which are solubilized by Cu/Zn-selective chelators in vitro. Here we report a 49% decrease in brain Abeta deposition (-375 microg/g wet weight, p = 0.0001) in a blinded study of APP2576 transgenic mice treated orally for 9 weeks with clioquinol, an antibiotic and bioavailable Cu/Zn chelator. This was accompanied by a modest increase in soluble Abeta (1.45% of total cerebral Abeta); APP, synaptophysin, and GFAP levels were unaffected. General health and body weight parameters were significantly more stable in the treated animals. These results support targeting the interactions of Cu and Zn with Abeta as a novel therapy for the prevention and treatment of AD.

Voltage-gated L-type Ca2+ channels (LTCCs) containing a pore-forming alpha1D subunit (D-LTCCs) are expressed in neurons and neuroendocrine cells. Their relative contribution to total L-type Ca2+ currents and their physiological role and significance as a drug target remain unknown. Therefore, we generated D-LTCC deficient mice (alpha1D-/-) that were viable with no major disturbances of glucose metabolism. alpha1D-/-mice were deaf due to the complete absence of L-type currents in cochlear inner hair cells and degeneration of outer and inner hair cells. In wild-type controls, D-LTCC-mediated currents showed low activation thresholds and slow inactivation kinetics. Electrocardiogram recordings revealed sinoatrial node dysfunction (bradycardia and arrhythmia) in alpha1D-/- mice. We conclude that alpha1D can form LTCCs with negative activation thresholds essential for normal auditory function and control of cardiac pacemaker activity.

One potential factor contributing to the susceptibility of these cells to premature death arises from the cytotoxic effects of amyloid-␤ (A␤) peptide deposition at or near sites of neuronal degeneration. Cultured human Franc ¸ois G.

Synthetic agonists of the growth hormone secretagogue receptor (GHSR) rejuvenate the pulsatile pattern of GH-release in the elderly, and increase lean but not fat mass in obese subjects. Screening of tissue extracts in a cell line engineered to overexpress the GHSR led to the identification of a natural agonist called ghrelin. Paradoxically, this hormone was linked to obesity. However, it had not been directly shown that the GHSR is a physiologically relevant ghrelin receptor. Furthermore, ghrelin's structure is significantly different from the synthetic agonist (MK-0677) used to expression-clone the GHSR. To address whether the GHSR mediates ghrelin's stimulatory effects on GH release and appetite, we generated Ghsr-null mice. In contrast to wild-type mice, acute treatment of Ghsr-null mice with ghrelin stimulated neither GH release nor food intake, showing that the GHSR is a biologically relevant ghrelin receptor. Nevertheless, Ghsr-null mice are not dwarfs; their appetite and body composition are comparable to that of wild-type littermates. Furthermore, in contrast to suggestions that ghrelin regulates leptin and insulin secretion, fastinginduced changes in serum levels of leptin and insulin are identical in wild-type and null mice. Serum insulin-like growth factor 1 levels and body weights of mature Ghsr-null mice are modestly reduced compared to wild-type littermates, which is consistent with ghrelin's property as an amplifier of GH pulsatility and its speculated role in establishing an insulin-like growth factor 1 set-point for maintaining anabolic metabolism. Our results suggest that chronic treatment with ghrelin antagonists will have little effect on growth or appetite.

Approximately 10% of cases of Alzheimer's disease are familial and associated with autosomal dominant inheritance of mutations in genes encoding the amyloid precursor protein, presenilin 1 (PS1) and presenilin 2 (PS2). Mutations in PS1 are linked to about 25% of cases of early-onset familial Alzheimer's disease. PS1, which is endoproteolytically processed in vivo, is a multipass transmembrane protein and is a functional homologue of SEL-12, a Caenorhabditis elegans protein that facilitates signalling mediated by the Notch/LIN-12 family of receptors. To examine potential roles for PS1 in facilitating Notch-mediated signalling during mammalian embryogenesis, we generated mice with targeted disruptions of PS1 alleles (PS1-/- mice). PS1-/- embryos exhibited abnormal patterning of the axial skeleton and spinal ganglia, phenotypes traced to defects in somite segmentation and differentiation. Moreover, expression of mRNA encoding Notch1 and Dll1 (delta-like gene 1), a vertebrate Notch ligand, is markedly reduced in the presomitic mesoderm of PS1-/- embryos compared to controls. Hence, PS1 is required for the spatiotemporal expression of Notch1 and Dll1, which are essential for somite segmentation and maintenance of somite borders.

In several pedigrees of early onset familial Alzheimer's disease (FAD), point mutations in the beta-amyloid precursor protein (APP) gene are genetically linked to the disease. This finding implicates APP in the pathogenesis of Alzheimer's disease in these individuals. To understand the in vivo function of APP and its processing, we have generated an APP-null mutation in mice. Homozygous APP-deficient mice were viable and fertile. However, the mutant animals weighed 15%-20% less than age-matched wild-type controls. Neurological evaluation showed that the APP-deficient mice exhibited a decreased locomotor activity and forelimb grip strength, indicating a compromised neuronal or muscular function. In addition, four out of six homozygous mice showed reactive gliosis at 14 weeks of age, suggesting an impaired neuronal function as a result of the APP-null mutation.

SUMMARY Abnormal NFκB activation has been implicated in Alzheimer’s disease (AD). However, the signaling pathways governing NFκB regulation and function in the brain are poorly understood. We identify complement protein C3 as an astroglial target of NFκB, and show that C3 release acts through neuronal C3aR to disrupt dendritic morphology and network function. Exposure to Aβ activates astroglial NFκB and C3 release, consistent with the high levels of C3 expression in brain tissue from AD patients and APP transgenic mice, where C3aR antagonist treatment rescues cognitive impairment. Thus, dysregulation of neuron-glia interaction through NFκB/C3/C3aR signaling may contribute to synaptic dysfunction in AD and C3aR antagonists may be therapeutically beneficial.

Notch signaling is a central mechanism for controlling embryogenesis. However, its in vivo function during mesenchymal cell differentiation, and specifically, in bone homeostasis remains largely unknown. Here, we show that osteoblast-specific gain of Notch function causes severe osteosclerosis due to increased proliferation of immature osteoblasts. Under these pathological conditions, Notch stimulates early osteoblastic proliferation by up-regulating Cyclin D, Cyclin E, and Osterix. Notch also regulates terminal osteoblastic differentiation by directly binding Runx2 and repressing its transactivation function. In contrast, loss of all physiologic Notch signaling in osteoblasts, generated by deletion of Presenilin 1 and 2 in bone, is associated with late onset, age-related osteoporosis resulting from increased osteoblast-dependent osteoclastic activity due to decreased production of Osteoprotegerin. Together, these findings highlight the potential dimorphic effects of Notch signaling in bone homeostasis and may provide direction for novel therapeutic applications.Evolutionarily conserved Notch signaling plays a critical role in cell fate determination, and various developmental processes by translating cell-cell interactions into specific transcriptional programs 1, 2 . Temporal and spatial modulation of this pathway can significantly affect proliferation, differentiation and apoptotic events 3 . Moreover, the timing of Notch signaling can lead to diverse effects within the same cell lineage 4, 5 . In mammals, activation of up to four Notch receptors by membrane-bound ligands initiates a process leading to presenilin-mediated cleavage and release of the Notch intracellular domain (NICD) from the membrane that then traffics to the nucleus. NICD subsequently regulates the expression of genes in cooperation with the transcription factor RBP-Jκ and Mastermind-like proteins.The observation that mutations in the Notch ligand Delta homologue-3 (Dll-3) and γ-secretase Presenilin1 both cause axial skeletal phenotypes originally linked Notch signaling with skeletal development 6, 7 . Recently, several in vitro studies with conflicting results implicated the Notch pathway in the regulation of osteoblast differentiation, but the in vivo role of Notch signaling in bone homeostasis still remains unknown 8-12 .Corresponding Author: Brendan Lee, M.D., Ph.D., One Baylor Plaza, Rm 635E, Houston, Tx 77030,, Email E-mail: blee@bcm.tmc.edu. In this study, we investigate the tissue, cellular, and molecular consequences of both gain and loss of function of Notch signaling in committed osteoblasts. NIH Public Access RESULTS Gain of function of Notch signaling results in severe osteosclerosisTo determine the pathological consequences of in vivo gain of Notch function during bone formation and homeostasis, we generated transgenic mice expressing the Notch1 intracellular domain (N1ICD) under the control of the type I collagen (Col1a1) promoter (Suppl. Fig. 1a,b). Here, gain of Notch function would occur in committed osteoblastic ce...