Surface expression and limited proteolysis of ADAM10 are increased by a dominant negative inhibitor of dynamin

<p>Abstract</p> <p>Background</p> <p>The amyloid precursor protein (APP) is cleaved by β- and γ-secretases to generate toxic amyloid β (Aβ) peptides. Alternatively, <it>α</it>-secretases cleave APP within the Aβ domain, precluding Aβ formation and releasing the soluble ectodomain, sAPPα. We previously showed that inhibition of the GTPase dynamin reduced APP internalization and increased release of sAPPα, apparently by prolonging the interaction between APP and α-secretases at the plasma membrane. This was accompanied by a reduction in Aβ generation. In the present study, we investigated whether surface expression of the α-secretase ADAM (a disintegrin and metalloprotease)10 is also regulated by dynamin-dependent endocytosis.</p> <p>Results</p> <p>Transfection of human embryonic kidney (HEK) cells stably expressing M3 muscarinic receptors with a dominant negative dynamin I mutant (dyn I K44A), increased surface expression of both immature, and mature, catalytically active forms of co-expressed ADAM10. Surface levels of ADAM10 were unaffected by activation of protein kinase C (PKC) or M3 receptors, indicating that receptor-coupled shedding of the ADAM substrate APP is unlikely to be mediated by inhibition of ADAM10 endocytosis in this cell line. Dyn I K44A strongly increased the formation of a C-terminal fragment of ADAM10, consistent with earlier reports that the ADAM10 ectodomain is itself a target for sheddases. The abundance of this fragment was increased in the presence of a γ-secretase inhibitor, but was not affected by M3 receptor activation. The dynamin mutant did not affect the distribution of ADAM10 and its C-terminal fragment between raft and non-raft membrane compartments.</p> <p>Conclusions</p> <p>Surface expression and limited proteolysis of ADAM10 are regulated by dynamin-dependent endocytosis, but are unaffected by activation of signaling pathways that upregulate shedding of ADAM substrates such as APP. Modulation of ADAM10 internalization could affect cellular behavior in two ways: by altering the putative signaling activity of the ADAM10 C-terminal fragment, and by regulating the biological function of ADAM10 substrates such as APP and N-cadherin.</p

Prenatal Choline Supplementation Protects against Postnatal Neurotoxicity

Choline, a dietary compound present in many foods, has recently been classified as an essential nutrient for humans. There is evidence from animal models that the availability of choline during the prenatal period influences neural and cognitive development. Here we report that choline supplementation during a 6 d gestational period protects against neurodegeneration in the posterior cingulate and retrosplenial cortices of female adolescent rats produced by peripheral administration of the NMDA receptor antagonist dizocilpine (MK-801). These data show that availability of a single nutrient, choline, during a brief period of prenatal development diminishes vulnerability to neurotoxicity in adolescent offspring

BMP9 Protects Septal Neurons from Axotomy-Evoked Loss of Cholinergic Phenotype

Cholinergic projection from the septum to the hippocampus is crucial for normal cognitive function and degeneration of cells and nerve fibers within the septohippocampal pathway contributes to the pathophysiology of Alzheimer's disease. Bone morphogenetic protein (BMP) 9 is a cholinergic differentiating factor during development both in vivo and in vitro.To determine whether BMP9 could protect the adult cholinergic septohippocampal pathway from axotomy-evoked loss of the cholinergic phenotype, we performed unilateral fimbria-fornix transection in mice and treated them with a continuous intracerebroventricular infusion of BMP9 for six days. The number of choline acetyltransferase (CHAT)-positive cells was reduced by 50% in the medial septal nucleus ipsilateral to the lesion as compared to the intact, contralateral side, and BMP9 infusion prevented this loss in a dose-dependent manner. Moreover, BMP9 prevented most of the decline of hippocampal acetylcholine levels ipsilateral to the lesion, and markedly increased CHAT, choline transporter CHT, NGF receptors p75 (NGFR-p75) and TrkA (NTRK1), and NGF protein content in both the lesioned and unlesioned hippocampi. In addition, BMP9 infusion reduced bilaterally hippocampal levels of basic FGF (FGF2) protein.These data indicate that BMP9 administration can prevent lesion-evoked impairment of the cholinergic septohippocampal neurons in adult mice and, by inducing NGF, establishes a trophic environment for these cells

MicroRNAs as Candidate Biomarkers for Alzheimer’s Disease

The neurological damage of Alzheimer’s disease (AD) is thought to be irreversible upon onset of dementia-like symptoms, as it takes years to decades for occult pathologic changes to become symptomatic. It is thus necessary to identify individuals at risk for the development of the disease before symptoms manifest in order to provide early intervention. Surrogate markers are critical for early disease detection, stratification of patients in clinical trials, prediction of disease progression, evaluation of response to treatment, and also insight into pathomechanisms. Here, we review the evidence for a number of microRNAs that may serve as biomarkers with possible mechanistic insights into the AD pathophysiologic processes, years before the clinical manifestation of the disease

THE JOURNAL OF PHARMACOLOGY ANI) EXPERIMENTAL THERAPEUTICS Formation of Methylamines from Ingested Choline and Lecithin1

ABSTRACT Human

Prenatal choline supplementation attenuates neuropathological response to status epilepticus in the adult rat hippocampus. Neurobiol. Dis

Prenatal choline supplementation (SUP) protects adult rats against spatial memory deficits observed after excitotoxin-induced status epilepticus (SE). To examine the mechanism underlying this neuroprotection, we determined the effects of SUP on a variety of hippocampal markers known to change in response to SE and thought to underlie ensuing cognitive deficits. Adult offspring from rat dams that received either a control or SUP diet on embryonic days 12-17 were administered saline or kainic acid (i.p.) to induce SE and were euthanized 16 days later. SUP markedly attenuated seizure-induced hippocampal neurodegeneration, dentate cell proliferation, and hippocampal GFAP mRNA expression levels, prevented the loss of hippocampal GAD65 protein and mRNA expression, and altered growth factor expression patterns. SUP also enhanced pre-seizure hippocampal levels of BDNF, NGF, and IGF-1, which may confer a neuroprotective hippocampal microenvironment that dampens the neuropathological response to and/or helps facilitate recovery from SE to protect cognitive function

BMP9 protects medial septum cholinergic neurons from axotomy-evoked degeneration.

<p>CHAT-positive cells were counted as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0021166#pone-0021166-g001" target="_blank">Fig. 1</a> on both sides of the medial septum and the average cell number on the lesioned side is expressed as % of the average cell number on the intact side. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0021166#pone-0021166-g002" target="_blank">Fig. 2</a> for representative sections. Using the best fit to a rectangular hyperbola (R² = 0.89) the EC₅₀ value for BMP9 was 1 ng/h. Note, at 15 ng/h and 38 ng/h BMP9 prevented all loss of CHAT-positive neurons.</p

FGF2 protein levels in hippocampus (HPP).

<p>Mice were treated as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0021166#pone-0021166-g004" target="_blank">Fig. 4</a>. FGF2 was measured by ELISA (* p<0.05 BMP9 vs PBS on the same side).</p

NGFR-p75, TrkA and NGF protein levels in hippocampus (HPP).

<p>Mice were treated as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0021166#pone-0021166-g004" target="_blank">Fig. 4</a>. NGFR-p75 (** p<0.01 BMP vs PBS on the same side) and TrkA (significant treatment effect of BMP9 by two-way ANOVA, p<0.05) were measured by Western blot (top panel) and the data quantified as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0021166#pone-0021166-g005" target="_blank">Fig. 5</a>. NGF was measured by ELISA (* p<0.05, ** p<0.01 BMP9 vs PBS on the same side).</p

Sections of mouse brain showing a representative image of the unilateral transection of fimbria-fornix (top panel) and the ROI stained with an anti-CHAT antibody (Bottom Panel).

<p>CHAT-positive cells are seen in the medial septum, the diagonal band and in the striatum (bottom panel). The lines show the ROI used for cell count analysis in the medial septum. The ROI was defined by a triangular shape that extended, dorso-ventrally, from the apex of the medial septum (A) to an imaginary line connecting the lower limits of the anterior commissures on each hemisphere (B) and, medio-laterally, from the midline (A) to the outer limits of the medial septal area (C and D).</p