Biomedical subjects

Daniel L Alkon

Publications and source records attributed to Daniel L Alkon.

At least 37 records · Page 2Linked to original sources

PubMed2004

Emerging targets for the pharmacology of learning and memory.

Learning and memory are dynamic processes associated with modifications in morphology, biochemistry and physiology of the nervous system, that can be analysed at different levels of biological organization. Within this context, changes correlated with cognitive processes are identifiable at distinct cellular and molecular loci of the nervous system. Synaptic plasticity represents an experience-dependent alteration of neuronal properties that subserve learning and memory, and for which a neuronal network can be considered as a candidate for being part of an engram. Multiple molecular systems at various levels of the signal transduction cascade, such as those involving receptors, second messengers and gene expression, may be linked to synaptic remodelling during cognitive processes. After a brief overview of the various facets of memory and the higher levels of cerebral organization (with special attention to the hippocampus), this review is devoted to the presentation of some novel potential therapeutic targets within the signal transduction cascade that might be considered for rational pharmacological intervention to improve learning and memory in both diseased and healthy individuals.

Animals↗

PubMed2004

Gene expression profiles of apoptotic neurons.

The multigenic program underlying neuronal apoptosis is mostly unknown. To study the program, we used genome-scale screening by oligonucleotide microarrays during serum and potassium deprivation-induced apoptosis of cerebellar granule neurons. From the 8740 genes interrogated by the arrays, 423 genes were found to be regulated at both the transcriptional and the posttranscriptional level and segregated into distinct clusters. Semantic clustering based on gene ontologies showed coordinated expression of genes with common biological functions and metabolic pathways. Among the genes implicated in apoptotic cerebellar granule neurons, 70 were in common with those differentially expressed in cortical neurons exposed to amyloid beta-protein, indicating the existence of common mechanisms responsible for neuronal cell death. Our results offer a genomic view of the changes that accompany neuronal apoptosis and yield new insights into the underlying molecular basis.

Amyloid beta-Peptides↗

PubMed2004

Two different biological configurations for long-term memory.

Long-term memory (LTM) in Hermissenda can be distinguished from consolidated long-term memory (CLTM) by determining how long recall is retained. LTM is retained for approximately 1 day, while CLTM is retained for at least 3 days. During the transition from LTM to CLTM, the extent of retention appears to depend partially on how much consolidation has been completed. Several models are discussed that may be related to the two different manifestations of recall.

Animals↗

PubMed2004

Cerebral hypoperfusion and amyloid-induced synergistic impairment of hippocampal CA1 synaptic efficacy and spatial memory in young adult rats.

Alzheimer's disease (AD) is characterized by amyloid plaques, neurofibrillary tangles in the brain, cerebral hypoperfusion/hypometabolism, and amyloid angiopathy. The former two and cell loss occur late in the disease and are probably not the leading causes for the initial memory decline. Cerebral hypoperfusion is a pre-clinical event in AD and represents the most accurate indicator predicting the probable AD patients to develop AD in a future time. However, in young animals, cerebral hypoperfusion as those matching the reduction in AD has no significant effects on learning and memory. Here, we report that association of cerebral hypoperfusion (2-vessel occlusion) with cerebrovascular amyloid (internal-carotid 0.5 mg beta(25-35), an active fragment of Abeta) significantly impaired spatial learning and memory of young adult rats, while neither the same insult alone had significant impact. At the time when the spatial memory was impaired, in vitro recording revealed that the associated cerebral hypoperfusion and internal-carotid amyloid reduced the ability of the hippocampal CA1 network to generate cholinergic theta and the synaptic modification evoked by associative activation of cholinergic and GABAergic inputs. The results suggest that cerebral hypoperfusion and amyloid angiopathy may play an important role as associated events in initiating the early memory decline in AD.

Alzheimer Disease↗

PubMed2003

Quantification and distribution of beta-secretase alternative splice variants in the rat and human brain.

Beta-amyloid (Abeta) is formed by sequential cleaving of the amyloid precursor protein by two proteolytic enzymes, beta- and gamma-secretases. Beta-secretase (BACE) is a type I transmembrane aspartic proteinase that is highly expressed in the mammalian brain. Four alternative splice variants of BACE are currently known and each encodes for a protein isoform with a different enzymatic activity. In Alzheimer's disease (AD) patients, the enzymatic activity and protein levels of BACE are increased in the neocortex, suggesting their differential expression may have a role in Abeta plaque formation. We have determined the differential expression of BACE mRNA and its splice variants in eight regions of the rat and two of the human brain. In humans, the frontal cortex which shows Abeta deposition in AD, expressed three-fold more BACE than the cerebellum and four fold more than the rats' frontal cortex both of which do not form Abeta plaques. The highest BACE levels of rats were found in the frontal cortex and less in other areas. Although most human and rat brain regions expressed all four BACE variants, the human cerebellum did not express the I-457 BACE variant. Human and rat frontal cortex expressed high levels of the I-501 and I-457 variants, but I-432 was highly expressed only in the rat. Species-specific differences were evident between human and rat brain areas, suggesting that BACE transcript variants may have different evolutionary conservation. Differential expression of BACE variants may explain the broad spectrum of phenotypic abnormalities and possible pathogenetic mechanisms underlying Alzheimer's disease.

Alternative Splicing↗

PubMed2003

Open space swimming test to index antidepressant activity.

Depression is characterized by a lack of "motivation" rather than a lack of "physical space" to move around. This study was designed to evaluate predictivity of an open space swimming test for antidepressant activity of various antidepressants in rats. Without drug treatment, rats showed a significant reduction in the distance moved (increased immobility) over successive trials in an open space water pool. Three major classes of antidepressants and a selective serotonin reuptake inhibitor (SSRI) were tested. Repeated treatment (10 mg/kg x 3 per day) of imipramine, a prototypical tricyclic antidepressant, iproniazid, a monoamine oxidase inhibitor, mianserin, an atypical antidepressant, and alaproclate, an SSRI, all significantly reduced the immobility. These results suggest that the open space swimming test is highly predictive of antidepressant action and is more sensitive to the drug treatments. The measurement is more objective than that of the forced swimming test and does not involve judging and scoring the animals' movement or lack of movement by investigators. The demonstrated effectiveness of three major types of antidepressants and an SSRI suggests that the effects on the test are not restricted to a particular underlying molecular mechanism of action. Thus, this swimming test shows promising potential as a screen for novel antidepressants and, perhaps, for revealing some of the underlying pathophysiology of depression.

Alanine↗

PubMed2003

c-Src protein tyrosine kinase activity is required for muscarinic receptor-mediated DNA synthesis and neurogenesis via ERK1/2 and c-AMP-responsive element-binding protein signaling in neural precursor cells.

The G protein-coupled muscarinic acetylcholine receptor (mAChR) isoforms have been identified in neural stem/progenitor (or precursor) cells. In previous studies, activation of these receptors induced elevations in intracellular Ca(2+) signals and mitogen-activated protein (MAP) kinase activity that led to enhanced DNA synthesis along with neurogenesis in neural precursor cells. Here we report that the nonreceptor protein tyrosine kinase c-src activity is required for the muscarinic receptor-activated MAP kinase and cAMP-responsive element-binding protein (CREB). Stimulation of neural precursor cells dissociated from embryonic day 13 rat cortical neuroepithelium with the muscarinic receptor agonist carbachol (CCh) induced phosphorylations of c-src that were detected by antibodies raised against phospho-Tyr416 (Ptyr416), phospho-Tyr527 (Ptyr527), and phospho-Tyr215 (Ptyr215) of the kinase. Although an increase in Ptyr416 suggested direct activation of c-src, Ptyr215 may serve as an alternative mechanism underlying activation of c-src without dephosphorylation of Ptyr-527. Both extracellular signal-regulated kinase (Erk1/2) and CREB were significantly activated after CCh treatment indicated by increases in phosphorylation of these two proteins. The c-Src inhibitor PP1 abolished the CCh-induced activation of Erk1/2 and CREB in a dose-dependent manner. Moreover, CCh stimulated expression of the neuronal specific marker MAP2, which was inhibited by PP1. Cell proliferation assays and immunocytochemistry revealed that PP1 inhibited the CCh-induced DNA synthesis and MAP2(+) production. These results suggest that c-src activity is essential for the muscarinic receptor-mediated proliferation and neurogenesis in neural precursor cells via Erk1/2 and CREB signaling pathways.

Animals↗

PubMed2003

Impairment of phosphatase 2A contributes to the prolonged MAP kinase phosphorylation in Alzheimer's disease fibroblasts.

The serine/threonine phosphatase 2A (PP2A) has been implicated in the pathogenesis of Alzheimer's disease (AD) due to its important role in regulating dephosphorylation of the microtubule-associated protein tau and mitogen-activated protein (MAP) kinase. In the present study, we show that PP2A was responsible for dephosphorylation of the extracellular signal-regulated kinase 1/2 (Erk1/2) following its activation by BK stimulation. Abnormal gene and protein expressions of PP2A, as well as its activity, were found to contribute to the abnormally prolonged Erk1/2 phosphorylation in the AD fibroblasts. Inhibition of PP2A with okadiac acid produced enhanced and more lasting Erk1/2 phosphorylation after BK stimulation, whereas FK506, an inhibitor of PP2B and FK-binding protein, inhibited the BK-stimulated Erk1/2 phosphorylation. Furthermore, while the phosphorylated Erk1/2 was concentrated in the nucleus of AC cells, it was mainly distributed in the extranuclear compartments of AD cells. These results suggest that the delayed dephosphorylation of Erk1/2 in AD cells following its BK-stimulated activation may be due to deficits of PP2A activity and impaired nuclear translocation of phosphorylated Erk1/2.

Active Transport, Cell Nucleus↗

PubMed2003

Time windows for effects of protein synthesis inhibitors on Pavlovian conditioning in Hermissenda: behavioral aspects.

Inhibitors of protein and mRNA syntheses inhibit long-term memory (LTM), but we lack information about the time windows during which those inhibitors are effective. Anisomycin (a protein synthesis translation inhibitor) was given to Hermissenda crassicornis which had received sufficient Pavlovian conditioning to produce LTM. When tested after 90 min, LTM recall was blocked when anisomycin was administered until 13 min after conditioning and from 65 to 75 min. There was good recall from 16 to 60 min. When tested at 240 min, two periods of sensitivity to anisomycin were revealed: one finished before 20 min, while the other lasted from 60 to 220 min.A brief study of the transcription inhibitor actinomycin-D found its action similar to that of anisomycin, except not inhibiting recall from about 160 to 220 min.

Animals↗

PubMed2003

Calcium-regulated GTPase activity in the calcium-binding protein calexcitin.

Calexcitin (CE) is a calcium-binding protein, closely related to sarcoplasmic calcium-binding proteins, that is involved in invertebrate learning and memory. Early reports indicated that both Hermissenda and squid CE also could bind GTP; however, the biochemical significance of GTP-binding and its relationship to calcium binding have remained unclear. Here, we report that the GTPase activity of CE is strongly regulated by calcium. CE possessed a P-loop-like structure near the C-terminal similar to the phosphate-binding regions in other GTP-binding proteins. Site-directed mutagenesis of this region showed that Gly(182), Phe(186) and Gly(187) are required for maximum affinity, suggesting that the GTP-binding motif is G-N-x-x-[FM]-G. CE cloned from Drosophila CNS possessed a similar C-terminal sequence and also bound and hydrolyzed GTP. GTPase activity in Drosophila CE was also strongly regulated by Ca(2+), exhibiting over 23-fold higher activity in the presence of 0.3 microM calcium. Analysis of the conserved protein motifs defines a new family of Ca(2+)-binding proteins representing the first example of proteins endowed with both EF-hand calcium binding domains and a C-terminal, P-loop-like GTP-binding motif. These results establish that, in the absence of calcium, both squid and Drosophila CE bind GTP at near-physiological concentrations and hydrolyze GTP at rates comparable to unactivated ras. Calcium functions to increase GTP-binding and GTPase activity in CE, similar to the effect of GTPase activating proteins in other low-MW GTP-binding proteins. CE may, therefore, act as a molecular interface between Ca(2+) cytosolic oscillations and the G protein-coupled signal transduction.

Amino Acid Sequence↗

PubMed2003

Gene expression of alpha-endosulfine in the rat brain: correlative changes with aging, learning and stress.

Endosulfine (EDSF) belongs to a highly conserved cAMP-regulated phosphoprotein (ARPP) family and was first isolated from ovine brain as a possible endogenous ligand for sulfonylurea receptors. To explore its involvement in brain functions, we investigated regional distribution of alpha-EDSF gene expression in the rat brain, and its regulation under physiological and pathological conditions. The majority of alpha-EDSF gene was expressed in the pyramidal neurons, which represent the principal excitatory neurons in various brain regions. Down-regulation of alpha-EDSF mRNA was detected in the rat hippocampus during long-term memory consolidation following a spatial learning experience, whereas swimming-related stress caused persistent up-regulation of alpha-EDSF gene expression in several brain regions. These changes, however, were absent from brains of diabetic rats that were subjected to the same behavioral treatments. Intracerebroventricular injection of streptozocin with a toxic dose induced severe learning deficits and brain structure alteration accompanied by a massive increase of alpha-EDSF mRNA in the somatosensory cortex. These results suggest that alpha-EDSF gene expression is differentially regulated by distinct brain processes involving excitatory neuronal activities.

Aging↗

PubMed2003

Regional rat brain distribution of heme oxygenase-1 and manganese superoxide dismutase mRNA: relevance of redox homeostasis in the aging processes.

Increasing evidence supports the notion that reduction of cellular expression and activity of antioxidant proteins and the resulting increase of oxidative stress are fundamental causes in the aging processes and neurodegenerative diseases. In the present study, we evaluated, in the brains of young and aged rats, the gene expression profiles of two inducible proteins critically involved in the cellular defense against endogenous or exogenous oxidants: heme oxygenase-1 (HO-1) and manganese superoxide dismutase-2 (SOD-2). SOD-2 is an essential antioxidant and HO-1 has been reported to be very active in regulating cellular redox homeostasis. Deregulation of these enzymes has been extensively reported to play a crucial role in the pathogenesis of neurodegenerative disorders. To measure the regional distribution of HO-1 and SOD-2 transcript levels in the rat brain, we have developed a real time quantitative reverse transcription-polymerase chain reaction protocol. Although these two genes presented a highly dissimilar range of expression, with SOD-2 >HO-1, both transcripts were highly expressed in the cerebellum and the hippocampus, showing in a different scale a strikingly parallel distribution gradient. To further investigate the regional brain expression of these mRNAs, we performed in situ hybridization using specific riboprobes. In situ hybridization results showed that both transcripts were highly concentrated in the hippocampus, the cerebellum and some specific regions of the brain cortex. We have also quantified, by reverse transcription-polymerase chain reaction, the brain expression of HO-1 and SOD-2 mRNAs in middle aged (12 months) and aged (28 months) rats. We found that the hippocampus of aged rats presents a significant down regulation of SOD2 mRNA expression and a parallel upregulation of HO-1 mRNA compared with young (6 months) and middle-aged rats. Furthermore, in the cerebellum of the aged rats, we detected a parallel significant upregulation of both HO-1 and SOD-2 transcripts. These regional age-dependent differences may help to explain the increased susceptibility to oxidative damage in these two brain areas during aging.

Aging↗

PubMed2002

Memory-specific temporal profiles of gene expression in the hippocampus.

Many experiments in the past have demonstrated the requirement of de novo gene expression during the long-term retention of learning and memory. Although previous studies implicated individual genes or genetic pathways in learning and memory, they did not uncover the collective behaviors or patterns of the genes. We have used genome-scale screening to analyze gene expression during spatial learning of rats in the Morris water maze. Our results show distinct temporal gene expression profiles associated with learning and memory. Exogenous administration of one peptide whose sustained increase during memory retention was implicated by microarray analysis, fibroblast growth factor (FGF)-18, improved spatial learning behavior, suggesting that pharmacological modulation of pathways and targets identified may allow new therapeutic approaches for improving learning and memory. Results of this study also suggest that while learning and physical activity involve common groups of genes, the behavior of learning and memory emerges from unique patterns of gene expression across time.

Animals↗

PubMed2002

Secretion of Annexin II via activation of insulin receptor and insulin-like growth factor receptor.

Annexin II is secreted into the extracellular environment, where, via interactions with specific proteases and extracellular matrix proteins, it participates in plasminogen activation, cell adhesion, and tumor metastasis and invasion. However, mechanisms regulating annexin II transport across the cellular membrane are unknown. In this study, we used coimmunoprecipitation to show that Annexin-II was bound to insulin and insulin-like growth factor-1 (IGF-1) receptors in PC12 cells and NIH-3T3 cells overexpressing insulin (NIH-3T3(IR)) or IGF-1 receptor (NIH-3T3(IGF-1R)). Stimulation of insulin and IGF-1 receptors by insulin caused a temporary dissociation of annexin II from these receptors, which was accompanied by an increased amount of extracellular annexin II detected in the media of PC12, NIH-3T3(IR), and NIH-3T3(IGF-1R) cells but not in that of untransfected NIH-3T3 cells. Activation of a different growth factor receptor, the platelet-derived growth factor receptor, did not produce such results. Tyrphostin AG1024, a tyrosine kinase inhibitor of insulin and IGF-1 receptor, was shown to inhibit annexin II secretion along with reduced receptor phosphorylation. Inhibitors of a few downstream signaling enzymes including phosphatidylinositol 3-kinase, pp60c-Src, and protein kinase C had no effect on insulin-induced annexin II secretion, suggesting a possible direct link between receptor activation and annexin II secretion. Immunocytochemistry revealed that insulin also induced transport of the membrane-bound form of annexin II to the outside layer of the cell membrane and appeared to promote cell aggregation. These results suggest that the insulin receptor and its signaling pathways may participate in molecular mechanisms mediating annexin II secretion.

3T3 Cells↗

PubMed2002

Gene expression profiles in a transgenic animal model of fragile X syndrome.

Fragile X syndrome is the most common inherited form of mental retardation. Although this syndrome originates from the absence of the RNA-binding protein FMRP, the molecular mechanisms underlying the cognitive deficits are unknown. The expression pattern of 6789 genes was studied in the brains of wild-type and FMR1 knockout mice, a fragile X syndrome animal model that has been associated with cognitive deficits. Differential expression of more than two-fold was observed for the brain mRNA levels of 73 genes. Differential expression of nine of these genes was confirmed by real-time quantitative reverse transcription-polymerase chain reaction and by in situ hybridization. In addition to corroborating the microarray data, the in situ hybridization analysis showed distinct spatial distribution patterns of microtubule-associated protein 2 and amyloid beta precursor protein. A number of differentially expressed genes associated with the fragile X syndrome phenotype have been previously involved in other memory or cognitive disorders.

Amyloid beta-Protein Precursor↗

PubMed2002

MAP kinase signaling cascade dysfunction specific to Alzheimer's disease in fibroblasts.

Mitogen-activated protein kinases (such as Erk1/2) regulate phosphorylation of the microtubule-associated protein tau and processing of the amyloid protein beta, both events critical to the pathophysiology of Alzheimer's disease (AD). Here we report that enhanced and prolonged Erk1/2 phosphorylation in response to bradykinin (BK) was detected in fibroblasts of both familial and sporadic AD, but not age-matched controls (AC). The AD-associated abnormality in Erk1/2 phosphorylation was not seen in fibroblasts from Huntington's disease patients with dementia. The elevation of Erk1/2 phosphorylation occurred immediately after BK stimulation and required an IP3-sensitive Ca(2+) release as well as activation of PKC and c-src as upstream events. Treatment of cells with the PI-3 kinase blocker LY924002 partially inhibited the BK-stimulated Erk1/2 phosphorylation in AC, but had no effect in AD cells, suggesting that the BK-induced Erk1/2 phosphorylation in AD cells is independent of PI-3 kinase. Activation of the cAMP-responsive element binding protein (CREB) monitored as an increase in phosphorylation at Ser-133 was also observed after BK stimulation. Unlike the AD-specific differences for Erk1/2, however, the BK-stimulated CREB phosphorylation was not different between AC and AD cells. Abnormal Erk1/2 activities may alter downstream cellular processes such as gene transcription, amyloid precursor protein processing, and tau protein phosphorylation, which contribute to the pathogenesis of AD. Moreover, detection of AD-specific differences in MAP kinase in peripheral tissues may provide an efficient means for early diagnosis of AD as well as help us to identify therapeutic targets for drug discovery.

Adult↗

PubMed2002

Carbonic anhydrase gating of attention: memory therapy and enhancement.

Enhancement of memory acquisition and recall represents an important pharmacological goal in the treatment of cognitive disorders. In addition to its involvement in pH regulation, HCO3- reabsorption and CO2 expiration, carbonic anhydrase plays a crucial role in signal processing, long-term synaptic transformation and attentional gating of memory storage. Carbonic anhydrase dysfunction impairs cognition and is associated with mental retardation, Alzheimer's disease and aging. The pharmacological profile of carbonic anhydrase has been refined and specific activators have been developed. In this article, an integrated view of the involvement of carbonic anhydrase activity in synaptic plasticity and cognition will be presented, with particular focus on attentional gating of spatial learning and memory.

Alzheimer Disease↗

PubMed2002

Programs of gene expression during the laying down of memory formation as revealed by DNA microarrays.

Many experiments in the past have demonstrated the requirement of de novo gene expression during memory formation. In contrast to the initial reductionistic view that genes relevant to learning and memory would be easily found and would provide a simple key to understand this brain function, it is becoming apparent that the genetic contribution to memory is complex. Previous approaches have been focused on individual genes or genetic pathways and failed to address the massively parallel nature of genome activities and collective behavior of the genes that ultimately control the molecular mechanisms underlying brain function. In view of the broad variety of genes and the cross talk of genetic pathways involved in this regulation, only gene expression profiles may reflect the complete behavior of regulatory pathways. In this review we illustrate how DNA microarray-based gene expression profiling may help to dissect and analyze the complex mechanisms involved in gene regulation during the acquisition and storage of memory in the mammalian brain.

Animals↗