Role of microtubule-dependent transport in synaptic plasticity in hippocampal neurons

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 12-12-2014The majority of excitatory synapses in the central nervous system are located at dendritic spines. These structures are considered key compartments for synaptic plasticity. During synaptic plasticity expression, spines alter their morphology and structure, and neurotransmitter receptor trafficking events take place. These phenomena have been related to actin cytoskeleton changes, but recently, microtubules (MT) have also been shown to modulate their own dynamics in an activity-dependent manner. Considering the close relationship between MT-dependent transport and endomembrane trafficking, and that between endomembrane trafficking and synaptic plasticity, we analyzed whether MTdependent transport would play a role in modulating synaptic plasticity. Using a multidisciplinary approach that includes biochemistry, live confocal imaging, electrophysiology techniques and molecular biology for the development of new molecular tools, we have investigated the interaction of AMPA Receptors (AMPARs) with members of the kinesin family (KIF5, KIF17) and dynein during synaptic function. In particular, we have mapped-down the domains of these molecular motors that are required for the maintenance of basal transmission and for the allowance of synaptic plasticity expression in CA1 excitatory synapses from hippocampal slices. The data obtained suggested that the motor domain of KIF5 is not needed either for the maintenance of basal transmission or for the transport of GluA2 subunits along dendrites. However, its motor activity, as well as the motor activity of KIF17, seems to be crucial for long-term depression (LTD) to occur normally. Complementarily to these results, we have found that maintenance of basal transmission most likely depends on either regulatory domains of the neck-stalk region, or on interactions depending on the cargo binding domain of KIF5 and KIF17. Finally, we have also addressed whether minus-end transport, based on dynein, could modulate synaptic function. Our preliminary results indicate that dynein transport is not required for proper synaptic function. Our results suggest that MT-dependent transport is one of the many intracellular mechanisms finely tuning NMDAR-dependent LTD. Altogehter, this thesis shed some light into the interplay between cytoskeletal elements and the regulation of synaptic strength.La mayoría de las sinapsis excitatorias en el sistema nervioso central se encuentran en espinas dendríticas. Por tanto, se consideran estructuras clave en las que la maquinaria necesaria para el desarrollo de la plasticidad sináptica debe funcionar correctamente. Cuando los fenómenos de plasticidad sináptica tienen lugar, las espinas modifican su morfología y estructura, y eventos de transporte de neurorreceptores tienen lugar. Estos procesos se han relacionado, clásicamente, con cambios en el citoesqueleto de actina, pero de un tiempo a esta parte se ha visto como los microtúbulos (MT) pueden alterar su dinámica en función de la actividad sináptica. Considerando, por una parte, la relación entre el transporte dependiente de MT y el tráfico endosomal; y, por otra, la relación entre el tráfico endosomal y los fenómenos de plasticidad sináptica; nos preguntamos explorar la posible relación entre el transporte mediado por MT y la plasticidad sináptica. Sirviéndonos de un enfoque multidisciplinar, en el que hemos utilizado técnicas de bioquímica, videomicroscopía confocal y electrofisiología, así como el desarrollo de nuevas herramientas moleculares; hemos investigado la interacción entre los receptores AMPA (AMPARs) y miembros de la familia de las kinesinas (KIF5, KIF17) y la dineina durante la expresión de la función sináptica. En concreto, hemos tratado de identificar los dominios de estos motores necesarios tanto para el mantenimiento de la transmisión basal, como para que tenga lugar una correcta plasticidad sináptica en sinapsis excitatorias de CA1 en rodajas de hipocampo. Así, hemos descubierto que, sorprendentemente, el dominio motor de KIF5 no es necesario ni para el mantenimiento de la transmisión basal, ni para el transporte de las subunidades GluA2 a lo largo de las dendritas. Sin embargo, su actividad motora, así como la de KIF17, sí parece crucial para una correcta expresión de la depresión a largo plazo (“long-term-depression” o LTD). De manera complementaria, hemos descubierto que el mantenimiento de la transmisión basal parece depender de dominios reguladores presentes en la región del cuello-tallo, o de interacciones mediadas por el dominio de unión a cargo de KIF5 o de KIF17. Por último, también hemos estudiado el posible papel del transporte en dirección menos, basado fundamentalmente en la actividad de la dineina. De manera preliminar, podemos concluir que el transporte en dirección menos no es necesario para el mantenimiento de la función sináptica. Nuestros resultados sugieren que la LTD dependiente de NMDA es un proceso altamente regulado por una gran variedad de mecanismos intracelulares, incluyendo el transporte mediado por MT. presentes en la región del cuello-tallo, o de interacciones mediadas por el dominio de unión a cargo de KIF5 o de KIF17. Por último, también hemos estudiado el posible papel del transporte en dirección menos, basado fundamentalmente en la actividad de la dineina. De manera preliminar, podemos concluir que el transporte en dirección menos no es necesario para el mantenimiento de la función sináptica. Nuestros resultados sugieren que la LTD dependiente de NMDA es un proceso altamente regulado por una gran variedad de mecanismos intracelulares, incluyendo el transporte mediado por MT

CSF tau microtubule-binding region identifies pathological changes in primary tauopathies

Despite recent advances in fluid biomarker research in Alzheimer's disease (AD), there are no fluid biomarkers or imaging tracers with utility for diagnosis and/or theragnosis available for other tauopathies. Using immunoprecipitation and mass spectrometry, we show that 4 repeat (4R) isoform-specific tau species from microtubule-binding region (MTBR-tau275 and MTBR-tau282) increase in the brains of corticobasal degeneration (CBD), progressive supranuclear palsy (PSP), frontotemporal lobar degeneration (FTLD)-MAPT and AD but decrease inversely in the cerebrospinal fluid (CSF) of CBD, FTLD-MAPT and AD compared to control and other FTLD-tau (for example, Pick's disease). CSF MTBR-tau measures are reproducible in repeated lumbar punctures and can be used to distinguish CBD from control (receiver operating characteristic area under the curve (AUC) = 0.889) and other FTLD-tau, such as PSP (AUC = 0.886). CSF MTBR-tau275 and MTBR-tau282 may represent the first affirmative biomarkers to aid in the diagnosis of primary tauopathies and facilitate clinical trial designs

Sex-specific effects of SNAP-25 genotype on verbal memory and Alzheimers disease biomarkers in clinically normal older adults.

INTRODUCTION: We tested sex-dependent associations of variation in the SNAP-25 gene, which encodes a presynaptic protein involved in hippocampal plasticity and memory, on cognitive and Alzheimers disease (AD) neuroimaging outcomes in clinically normal adults. METHODS: Participants were genotyped for SNAP-25 rs1051312 (T > C; SNAP-25 expression: C-allele > T/T). In a discovery cohort (N = 311), we tested the sex by SNAP-25 variant interaction on cognition, Aβ-PET positivity, and temporal lobe volumes. Cognitive models were replicated in an independent cohort (N = 82). RESULTS: In the discovery cohort, C-allele carriers exhibited better verbal memory and language, lower Aβ-PET positivity rates, and larger temporal volumes than T/T homozygotes among females, but not males. Larger temporal volumes related to better verbal memory only in C-carrier females. The female-specific C-allele verbal memory advantage was evidenced in the replication cohort. CONCLUSIONS: In females, genetic variation in SNAP-25 is associated with resistance to amyloid plaque formation and may support verbal memory through fortification of temporal lobe architecture. HIGHLIGHTS: The SNAP-25 rs1051312 (T > C) C-allele results in higher basal SNAP-25 expression. C-allele carriers had better verbal memory in clinically normal women, but not men. Female C-carriers had higher temporal lobe volumes, which predicted verbal memory. Female C-carriers also exhibited the lowest rates of amyloid-beta PET positivity. The SNAP-25 gene may influence female-specific resistance to Alzheimers disease (AD)

Gamma neuromodulation improves episodic memory and its associated network in amnestic mild cognitive impairment: a pilot study

Amnestic mild cognitive impairment (aMCI) is a predementia stage of Alzheimer's disease associated with dysfunctional episodic memory and limited treatment options. We aimed to characterize feasibility, clinical, and biomarker effects of noninvasive neurostimulation for aMCI. 13 individuals with aMCI received eight 60-minute sessions of 40-Hz (gamma) transcranial alternating current stimulation (tACS) targeting regions related to episodic memory processing. Feasibility, episodic memory, and plasma Alzheimer's disease biomarkers were assessed. Neuroplastic changes were characterized by resting-state functional connectivity (RSFC) and neuronal excitatory/inhibitory balance. Gamma tACS was feasible and aMCI participants demonstrated improvement in multiple metrics of episodic memory, but no changes in biomarkers. Improvements in episodic memory were most pronounced in participants who had the highest modeled tACS-induced electric fields and exhibited the greatest changes in RSFC. Increased RSFC was also associated with greater hippocampal excitability and higher baseline white matter integrity. This study highlights initial feasibility and the potential of gamma tACS to rescue episodic memory in an aMCI population by modulating connectivity and excitability within an episodic memory network

Hepatic and renal function impact concentrations of plasma biomarkers of neuropathology.

IntroductionThe impact of hepatorenal function on plasma biomarkers of neuropathology is unknown. Herein, we measured several plasma biomarkers in patients with cirrhosis.MethodsPlasma phosphorylated tau (p-tau181), neurofilament light chain (NfL), glial fibrillary acidic protein (GFAP), total tau (t-tau), and ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) were measured in 135 adults with cirrhosis and 22 healthy controls using Simoa. Within cirrhosis, associations between biomarkers and hepatorenal function were explored using linear regression.Resultsp-tau181, NfL, t-tau, and UCHL1 were increased 2- to 4-fold in cirrhosis, whereas GFAP was not increased. Within cirrhosis, creatinine moderately correlated with p-tau181 (β = 0.75, P < .01), NfL (β = 0.32, P < .01), and t-tau (β = 0.31, P < .01), but not GFAP (β = -0.01, P = .88) or UCHL1 (β = -0.05, P = .60), whereas albumin showed weak, inverse correlations: p-tau181 (β = -0.18, P < .01), NfL (β = -0.22, P < .01), GFAP (β = -0.17, P < .05), t-tau (β = -0.20, P = .02), and UCHL1 (β = -0.15, P = .09).ConclusionsElevated p-tau181, NfL, and t-tau in cirrhosis were associated with renal impairment and hypoalbuminemia, suggesting that hepatorenal function may be important when interpreting plasma biomarkers of neuropathology

Clinicopathological correlation of cerebrospinal fluid alpha‐synuclein seed amplification assay in a behavioral neurology autopsy cohort

IntroductionLewy body disease (LBD) is a common primary or co-pathology in neurodegenerative syndromes. An alpha-synuclein seed amplification assay (αSyn-SAA) is clinically available, but clinical performance, especially lower sensitivity in amygdala-predominant cases, is not well understood.MethodsAntemortem CSF from neuropathology-confirmed LBD cases was tested with αSyn-SAA (N = 56). Diagnostic performance and clinicopathological correlations were examined.ResultsSimilar to prior reports, sensitivity was 100% for diffuse and transitional LBD (9/9), and overall specificity was 96.3% (26/27). Sensitivity was lower in amygdala-predominant (6/14, 42.8%) and brainstem-predominant LBD (1/6, 16.7%), but early spread outside these regions (without meeting criteria for higher stage) was more common in αSyn-SAA-positive cases (6/7, 85.7%) than negative (2/13, 15.4%).DiscussionIn this behavioral neurology cohort, αSyn-SAA had excellent diagnostic performance for cortical LBD. In amygdala- and brainstem-predominant cases, sensitivity was lower, but positivity was associated with anatomical spread, suggesting αSyn-SAA detects early LBD progression in these cohorts.HighlightsA cerebrospinal fluid alpha-synuclein assay detects cortical LBD with high sensitivity/specificity. Positivity in prodromal stages of LBD was associated with early cortical spread. The assay provides precision diagnosis of LBD that could support clinical trials. The assay can also identify LBD co-pathology, which may impact treatment responses

Plasma neurofilament light chain levels suggest neuroaxonal stability following therapeutic remyelination in people with multiple sclerosis

Background Chronic demyelination is a major contributor to axonal vulnerability in multiple sclerosis (MS). Therefore, remyelination could provide a potent neuroprotective strategy. The ReBUILD trial was the first study showing evidence for successful remyelination following treatment with clemastine in people with MS (pwMS) with no evidence of disease activity or progression (NEDAP). Whether remyelination was associated with neuroprotection remains unexplored.Methods Plasma neurofilament light chain (NfL) levels were measured from ReBUILD trial's participants. Mixed linear effect models were fit for individual patients, epoch and longitudinal measurements to compare NfL concentrations between samples collected during the active and placebo treatment period.Results NfL concentrations were 9.6% lower in samples collected during the active treatment with clemastine (n=53, geometric mean=6.33 pg/mL) compared to samples collected during treatment with placebo (n=73, 7.00 pg/mL) (B=-0.035 [-0.068 to -0.001], p=0.041). Applying age- and body mass index-standardised NfL Z-scores and percentiles revealed similar results (0.04 vs 0.35, and 27.5 vs 33.3, p=0.023 and 0.042, respectively). Higher NfL concentrations were associated with more delayed P100 latencies (B=1.33 [0.26 to 2.41], p=0.015). In addition, improvement of P100 latencies between visits was associated with a trend for lower NfL values (B=0.003 [-0.0004 to 0.007], p=0.081). Based on a Cohen's d of 0.248, a future 1:1 parallel-arm placebo-controlled study using a remyelinating agent with comparable effect as clemastine would need 202 subjects per group to achieve 80% power.Conclusions In pwMS, treatment with the remyelinating agent clemastine was associated with a reduction of blood NfL, suggesting that neuroprotection is achievable and measurable with therapeutic remyelination

Presence of Preoperative Neurodegeneration Biofluid Markers in Patients with Postoperative Delirium

BackgroundThe pathophysiology of delirium is incompletely understood, including what molecular pathways are involved in brain vulnerability to delirium. This study examined whether preoperative plasma neurodegeneration markers were elevated in patients who subsequently developed postoperative delirium through a retrospective case-control study.MethodsInclusion criteria were patients of 65 yr of age or older, undergoing elective noncardiac surgery with a hospital stay of 2 days or more. Concentrations of preoperative plasma P-Tau181, neurofilament light chain, amyloid β1-42 (Aβ42), and glial fibrillary acidic protein were measured with a digital immunoassay platform. The primary outcome was postoperative delirium measured by the Confusion Assessment Method. The study included propensity score matching by age and sex with nearest neighbor, such that each patient in the delirium group was matched by age and sex with a patient in the no-delirium group.ResultsThe initial cohort consists of 189 patients with no delirium and 102 patients who developed postoperative delirium. Of 291 patients aged 72.5 ± 5.8 yr, 50.5% were women, and 102 (35%) developed postoperative delirium. The final cohort in the analysis consisted of a no-delirium group (n = 102) and a delirium group (n = 102) matched by age and sex using the propensity score method. Of the four biomarkers assayed, the median value for neurofilament light chain was 32.05 pg/ml for the delirium group versus 23.7 pg/ml in the no-delirium group. The distribution of biomarker values significantly differed between the delirium and no-delirium groups (P = 0.02 by the Kolmogorov-Smirnov test) with the largest cumulative probability difference appearing at the biomarker value of 32.05 pg/ml.ConclusionsThese results suggest that patients who subsequently developed delirium are more likely to be experiencing clinically silent neurodegenerative changes before surgery, reflected by changes in plasma neurofilament light chain biomarker concentrations, which may identify individuals with a preoperative vulnerability to subsequent cognitive decline.Editor’s perspectiv

Scalable plasma and digital cognitive markers for diagnosis and prognosis of Alzheimer's disease and related dementias

IntroductionWith emergence of disease-modifying therapies, efficient diagnostic pathways are critically needed to identify treatment candidates, evaluate disease severity, and support prognosis. A combination of plasma biomarkers and brief digital cognitive assessments could provide a scalable alternative to current diagnostic work-up.MethodsWe examined the accuracy of plasma biomarkers and a 10-minute supervised tablet-based cognitive assessment (Tablet-based Cognitive Assessment Tool Brain Health Assessment [TabCAT-BHA]) in predicting amyloid β positive (Aβ+) status on positron emission tomography (PET), concurrent disease severity, and functional decline in 309 older adults with subjective cognitive impairment (n = 49), mild cognitive impairment (n = 159), and dementia (n = 101).ResultsCombination of plasma pTau181, Aβ42/40, neurofilament light (NfL), and TabCAT-BHA was optimal for predicting Aβ-PET positivity (AUC = 0.962). Whereas NfL and TabCAT-BHA optimally predicted concurrent disease severity, combining these with pTau181 and glial fibrillary acidic protein was most accurate in predicting functional decline.DiscussionCombinations of plasma and digital cognitive markers show promise for scalable diagnosis and prognosis of ADRD.HighlightsThe need for cost-efficient diagnostic and prognostic markers of AD is urgent. Plasma and digital cognitive markers provide complementary diagnostic contributions. Combination of these markers holds promise for scalable diagnosis and prognosis. Future validation in community cohorts is needed to inform clinical implementation

Plasma phosphorylated tau 217 and phosphorylated tau 181 as biomarkers in Alzheimer's disease and frontotemporal lobar degeneration : a retrospective diagnostic performance study

Background: Plasma tau phosphorylated at threonine 217 (p-tau217) and plasma tau phosphorylated at threonine 181 (p-tau181) are associated with Alzheimer's disease tau pathology. We compared the diagnostic value of both biomarkers in cognitively unimpaired participants and patients with a clinical diagnosis of mild cognitive impairment, Alzheimer's disease syndromes, or frontotemporal lobar degeneration (FTLD) syndromes. Methods: In this retrospective multicohort diagnostic performance study, we analysed plasma samples, obtained from patients aged 18–99 years old who had been diagnosed with Alzheimer's disease syndromes (Alzheimer's disease dementia, logopenic variant primary progressive aphasia, or posterior cortical atrophy), FTLD syndromes (corticobasal syndrome, progressive supranuclear palsy, behavioural variant frontotemporal dementia, non-fluent variant primary progressive aphasia, or semantic variant primary progressive aphasia), or mild cognitive impairment; the participants were from the University of California San Francisco (UCSF) Memory and Aging Center, San Francisco, CA, USA, and the Advancing Research and Treatment for Frontotemporal Lobar Degeneration Consortium (ARTFL; 17 sites in the USA and two in Canada). Participants from both cohorts were carefully characterised, including assessments of CSF p-tau181, amyloid-PET or tau-PET (or both), and clinical and cognitive evaluations. Plasma p-tau181 and p-tau217 were measured using electrochemiluminescence-based assays, which differed only in the biotinylated antibody epitope specificity. Receiver operating characteristic analyses were used to determine diagnostic accuracy of both plasma markers using clinical diagnosis, neuropathological findings, and amyloid-PET and tau-PET measures as gold standards. Difference between two area under the curve (AUC) analyses were tested with the Delong test. Findings: Data were collected from 593 participants (443 from UCSF and 150 from ARTFL, mean age 64 years [SD 13], 294 [50%] women) between July 1 and Nov 30, 2020. Plasma p-tau217 and p-tau181 were correlated (r=0·90, p<0·0001). Both p-tau217 and p-tau181 concentrations were increased in people with Alzheimer's disease syndromes (n=75, mean age 65 years [SD 10]) relative to cognitively unimpaired controls (n=118, mean age 61 years [SD 18]; AUC=0·98 [95% CI 0·95–1·00] for p-tau217, AUC=0·97 [0·94–0·99] for p-tau181; pdiff=0·31) and in pathology-confirmed Alzheimer's disease (n=15, mean age 73 years [SD 12]) versus pathologically confirmed FTLD (n=68, mean age 67 years [SD 8]; AUC=0·96 [0·92–1·00] for p-tau217, AUC=0·91 [0·82–1·00] for p-tau181; pdiff=0·22). P-tau217 outperformed p-tau181 in differentiating patients with Alzheimer's disease syndromes (n=75) from those with FTLD syndromes (n=274, mean age 67 years [SD 9]; AUC=0·93 [0·91–0·96] for p-tau217, AUC=0·91 [0·88–0·94] for p-tau181; pdiff=0·01). P-tau217 was a stronger indicator of amyloid-PET positivity (n=146, AUC=0·91 [0·88–0·94]) than was p-tau181 (n=214, AUC=0·89 [0·86–0·93]; pdiff=0·049). Tau-PET binding in the temporal cortex was more strongly associated with p-tau217 than p-tau181 (r=0·80 vs r=0·72; pdiff<0·0001, n=230). Interpretation: Both p-tau217 and p-tau181 had excellent diagnostic performance for differentiating patients with Alzheimer's disease syndromes from other neurodegenerative disorders. There was some evidence in favour of p-tau217 compared with p-tau181 for differential diagnosis of Alzheimer's disease syndromes versus FTLD syndromes, as an indication of amyloid-PET-positivity, and for stronger correlations with tau-PET signal. Pending replication in independent, diverse, and older cohorts, plasma p-tau217 and p-tau181 could be useful screening tools to identify individuals with underlying amyloid and Alzheimer's disease tau pathology. Funding: US National Institutes of Health, State of California Department of Health Services, Rainwater Charitable Foundation, Michael J Fox foundation, Association for Frontotemporal Degeneration, Alzheimer's Association