Robert G. Struble

Nicotinic cholinoceptive neurons of the frontal cortex are reduced in Alzheimer's disease

The cellular distribution of nicotinic acetylcholine receptors was studied in the frontal cortex (area 10) of 1) Alzheimer patients and compared to 2) age-matched and 3) middle-aged controls using the monoclonal antibody WF 6 and an immunoperoxidase protocol. Statistical analysis revealed significant differences between the number of labeled neurons among all three groups tested (middle-aged controls greater than aged controls greater than Alzheimer cases). No differences were seen for cresyl violet-stained samples. These findings underline that the nicotinic receptor decrease found with radioligand binding may reflect a postsynaptic in addition to a presynaptic component.

Apolipoprotein E4 inhibits, and apolipoprotein E3 promotes neurite outgrowth in cultured adult mouse cortical neurons through the low-density lipoprotein receptor-related protein

The apolipoprotein E4 (apoE4) genotype is a major risk factor for Alzheimers disease (AD); however, the mechanism is unknown. We previously demonstrated that apoE isoforms differentially modulated neurite outgrowth in embryonic neurons and in neuronal cell lines. ApoE3 increased neurite outgrowth whereas apoE4 decreased outgrowth, suggesting that apoE4 may directly affect neurons in the brain. In the present study we examined the effects of apoE on neurite outgrowth from cultured adult mouse cortical neurons to examine if adult neurons respond the same way that embryonic cells do. The results from this study demonstrated that (1) cortical neurons derived from adult apoE-gene knockout (apoE KO) mice have significantly shorter neurites than neurons from adult wild-type (WT) mice; (2) incubation of cortical neurons from adult apoE KO mice with human apoE3 increased neurite outgrowth, whereas human apoE4 decreased outgrowth in a dose-dependent fashion; (3) the isoform specific effects were abolished by incubation of the neurons with either receptor associated protein (RAP) or lactoferrin, both of which block the interaction of apoE-containing lipoproteins with the low-density lipoprotein receptor-related protein (LRP). These data suggest a potential mechanism whereby apoE4 may play a role in regenerative failure and accelerate the development of AD.

β-Secretase-1 elevation in transgenic mouse models of Alzheimer's disease is associated with synaptic/axonal pathology and amyloidogenesis: implications for neuritic plaque development

The presence of neuritic plaques is a pathological hallmark of Alzheimer’s disease (AD). However, the origin of extracellular β‐amyloid peptide (Aβ) deposits and the process of plaque development remain poorly understood. The present study attempted to explore plaque pathogenesis by localizing β‐secretase‐1 (BACE1) elevation relative to Aβ accumulation and synaptic/neuritic alterations in the forebrain, using transgenic mice harboring familial AD (FAD) mutations (5XFAD and 2XFAD) as models. In animals with fully developed plaque pathology, locally elevated BACE1 immunoreactivity (IR) coexisted with compact‐like Aβ deposition, with BACE1 IR occurring selectively in dystrophic axons of various neuronal phenotypes or origins (GABAergic, glutamatergic, cholinergic or catecholaminergic). Prior to plaque onset, localized BACE1/Aβ IR occurred at swollen presynaptic terminals and fine axonal processes. These BACE1/Aβ‐containing axonal elements appeared to undergo a continuing process of sprouting/swelling and dystrophy, during which extracellular Aβ IR emerged and accumulated in surrounding extracellular space. These data suggest that BACE1 elevation and associated Aβ overproduction inside the sprouting/dystrophic axonal terminals coincide with the onset and accumulation of extracellular amyloid deposition during the development of neuritic plaques in transgenic models of AD. Our findings appear to be in harmony with an early hypothesis that axonal pathogenesis plays a key or leading role in plaque formation.

Doublecortin expression in adult cat and primate cerebral cortex relates to immature neurons that develop into GABAergic subgroups

DCX-immunoreactive (DCX+) cells occur in the piriform cortex in adult mice and rats, but also in the neocortex in adult guinea pigs and rabbits. Here we describe these cells in adult domestic cats and primates. In cats and rhesus monkeys, DCX+ cells existed across the allo- and neocortex, with an overall ventrodorsal high to low gradient at a given frontal plane. Labeled cells formed a cellular band in layers II and upper III, exhibiting dramatic differences in somal size (5-20 microm), shape (unipolar, bipolar, multipolar and irregular), neuritic complexity and labeling intensity. Cell clusters were also seen in this band, and those in the entorhinal cortex extended into deeper layers as chain-like structures. Densitometry revealed a parallel decline of the cells across regions with age in cats. Besides the cellular band, medium-sized cells with weak DCX reactivity resided sparsely in other layers. Throughout the cortex, virtually all DCX+ cells co-expressed polysialylated neural cell adhesion molecule. Medium to large mature-looking DCX+ cells frequently colocalized with neuron-specific nuclear protein and gamma-aminobutyric acid (GABA), and those with a reduced DCX expression also partially co-labeled for glutamic acid decarboxylase, parvalbumin, calbindin, beta-nicotinamide adenine dinucleotide phosphate diaphorase and neuronal nitric oxide synthase. Similar to cats and monkeys, small and larger DCX+ cells were detected in surgically removed human frontal and temporal cortices. These data suggest that immature neurons persist into adulthood in many cortical areas in cats and primates, and that these cells appear to undergo development and differentiation to become functional subgroups of GABAergic interneurons.

Olfactory bulb lesions in alzheimer's disease

The olfactory bulb (OB), with its comparatively simple and well-delineated connectivity, presents an interesting system for examining cell-specific pathology in neurologic degenerative disorders such as Alzheimers disease (AD). We have found that in AD the large, efferently projecting neurons (mitral cells) of the OB degenerate, typically without classical Alzheimer neurofibrillary changes. In some cases, with less severe neocortical pathology, the terminal arborizations of olfactory nerve appear hyperplastic and are associated with focal accumulations of A-4 (beta-amyloid) immunoreactivity that are not detectable by standard amyloid stains. These abnormalities may represent a pathologic manifestation of normally occurring plasticity in the olfactory system.

Is Brain Amyloid Production a Cause or a Result of Dementia of The Alzheimer's Type?

The amyloid cascade hypothesis has guided much of the research into Alzheimers disease (AD) over the last 25 years. We argue that the hypothesis of amyloid-β (Aβ) as the primary cause of dementia may not be fully correct. Rather, we propose that decline in brain metabolic activity, which is tightly linked to synaptic activity, actually underlies both the cognitive decline in AD and the deposition of Aβ. Aβ may further exacerbate metabolic decline and result in a downward spiral of cognitive function, leading to dementia. This novel interpretation can tie the disparate risk factors for dementia to a unifying hypothesis and present a roadmap for interventions to decrease the prevalence of dementia in the elderly population.

Olfactory function in apoE knockout mice.

Apolipoprotein E (apoE), a lipid transporting protein, has been shown to play a vital role in nerve repair and remodeling. Since the olfactory system is in a continuous state of remodeling, the present study tested the hypothesis that apoE is required for normal functioning of the olfactory system. Olfactory behavior of wild-type (WT) and apoE-deficient (apoE KO) mice was assessed by using three standard olfactory tests: (1) the buried food pellet (BFP) test; (2) the odor choice (OC) test; and (3) the odor cued taste avoidance (OCTA) test. ApoE KO mice performed poorly in all the three tests as compared to WT mice, although they learned the tasks at a rate comparable to WT mice. ApoE KO mice had a significantly longer latency to find the buried pellet than WT mice. In the OC experiment, apoE KO mice did not differentiate water from an odorant solution. Furthermore, in the OCTA test the apoE KO mice were significantly less successful than WT mice at avoiding water containing an odorant and a bad tastant. These data demonstrate that apoE deficiency in apoE KO mice leads to a deficit in olfactory function, suggesting an important role for apoE in the olfactory system.

β-Secretase-1 elevation in aged monkey and Alzheimer’s disease human cerebral cortex occurs around the vasculature in partnership with multisystem axon terminal pathogenesis and β-amyloid accumulation

Alzheimer’s disease (AD) is the most common dementia‐causing disorder in the elderly; it may be related to multiple risk factors, and is characterized pathologically by cerebral hypometabolism, paravascular β‐amyloid peptide (Aβ) plaques, neuritic dystrophy, and intra‐neuronal aggregation of phosphorylated tau. To explore potential pathogenic links among some of these lesions, we examined β‐secretase‐1 (BACE1) alterations relative to Aβ deposition, neuritic pathology and vascular organization in aged monkey and AD human cerebral cortex. Western blot analyses detected increased levels of BACE1 protein and β‐site‐cleavage amyloid precursor protein C‐terminal fragments in plaque‐bearing human and monkey cortex relative to controls. In immunohistochemistry, locally elevated BACE1 immunoreactivity (IR) occurred in AD but not in control human cortex, with a trend for increased overall density among cases with greater plaque pathology. In double‐labeling preparations, BACE1 IR colocalized with immunolabeling for Aβ but not for phosphorylated tau. In perfusion‐fixed monkey cortex, locally increased BACE1 IR co‐existed with intra‐axonal and extracellular Aβ IR among virtually all neuritic plaques, ranging from primitive to typical cored forms. This BACE1 labeling localized to swollen/sprouting axon terminals that might co‐express one or another neuronal phenotype markers (GABAergic, glutamatergic, cholinergic, or catecholaminergic). Importantly, these BACE1‐labeled dystrophic axons resided near to or in direct contact with blood vessels. These findings suggest that plaque formation in AD or normal aged primates relates to a multisystem axonal pathogenesis that occurs in partnership with a potential vascular or metabolic deficit. The data provide a mechanistic explanation for why senile plaques are present preferentially near the cerebral vasculature.

Estradiol regulation of astroglia and apolipoprotein E: an important role in neuronal regeneration.

The effects of ovarian hormone on neuronal growth and function are well known. However, equally important, but often neglected, are ovarian hormone effects on glia. Our in vivo and in vitro studies show that estradiol modifies both neuronal growth and glial activity and these effects are tightly linked. Estradiol stimulates neurite growth and the release of the glial apolipoprotein E (apoE) in culture studies. Estradiol-stimulated neurite growth in these cultures requires apoE. Estradiol replacement in ovariectomized mice transiently increases the expression of apoE, the low density lipoprotein receptor related protein (LRP) and synaptophysin throughout the brain. Continuous estradiol replacement over two months loses effect on apoE, LRP, and synaptophysin and suppresses reactive gliosis. Estrous cycle variation of glial activation (GFAP) and apoE are not identical. We propose that estradiol (and other ovarian hormones) functions as a zeitgeber to co-ordinate neuronal-glial interactions. Co-ordination assures temporally appropriate excitatory and inhibitory interactions between glia and neurons. With aging and the loss of ovarian cyclicity, some of this co-ordination must be diminished. These observations present significant clinical implications. Approaches to hormone therapy (HT), for diminishing the risk of chronic neurological diseases, need to consider the temporal nature of ovarian hormones in brain repair and plasticity. Moreover, approaches must consider apoE genotype. The neuroprotective effects of HT in numerous chronic age-related diseases may represent effective co-ordination of repair processes rather than direct disease-specific actions. Moreover, the role of glial-derived proteins in neuroprotection should not be ignored.

Time course of response to estradiol replacement in ovariectomized mice: brain apolipoprotein E and synaptophysin transiently increase and glial fibrillary acidic protein is suppressed.

The current study examined the effect of long-term estradiol replacement in ovariectomized mice. Estradiol-17beta (E2) pellets or vehicle pellets were implanted at the time of ovariectomy (OVX) in young adult female mice. Five mice from each group were sacrificed at 5, 14, 28 and 49 days after OVX and pellet replacement. Western blotting of homogenates from somatosensory cortex, hippocampus, olfactory bulb and cerebellum was performed to obtain concentrations of glial fibrillary acidic protein (GFAP), apolipoprotein E (apoE) and synaptophysin (SYN). At 5 days after OVX, GFAP levels were not affected by E2 replacement. In contrast to GFAP, synaptophysin and apoE concentrations were significantly elevated by 15% and 25%, respectively, in the E2-replaced group compared to the vehicle-replaced group at 5 days but by 14 days concentrations were equivalent. Late in the time course of this study, at 49 days, GFAP concentrations were higher in the E2-deprived mice but did not increase in the E2-replaced group. Immunocytochemistry for GFAP confirmed this observation. Of note was that these effects occurred in all four brain regions measured. These observations suggest that estradiol is able to suppress reactive gliosis. In addition, E2 replacement in OVX mice is associated with transiently higher levels of apoE and synaptophysin.