Manuel Sarasa

Role of thyroid hormone in regulation of renal phosphate transport in young and aged rats.

In the present study, we have examined the cellular mechanisms mediating the regulation of renal proximal tubular sodium-coupled inorganic phosphate (Na/Pi) transport by thyroid hormone (T3) in young and aged rats. Young hypothyroid rats showed a marked decrease in Na/Pi cotransport activity, which was associated with parallel decreases in type II Na/Pi cotransporter (NaPi-2) protein and messenger RNA (mRNA) abundance. In contrast, administration of long-term physiological and supraphysiological doses of T3 resulted in significant increases in Na/Pi cotransport activity, protein, and mRNA levels. Nuclear run-on experiments indicated that thyroid hormone regulates NaPi-2 mRNA levels by a transcriptional mechanism. In aged rats, although there were no changes in T3 serum levels (when compared with young animals), there were significant decreases in serum Pi concentration, renal Na/Pi cotransport activity, and NaPi-2 protein and mRNA abundance. These effects were mediated, at least in part, by a reduction in...

Natural Non-Trasgenic Animal Models for Research in Alzheimer’s Disease

The most common animal models currently used for Alzheimer disease (AD) research are transgenic mice that express a mutant form of human Aβ precursor protein (APP) and/or some of the enzymes implicated in their metabolic processing. However, these transgenic mice carry their own APP and APP-processing enzymes, which may interfere in the production of different amyloid-beta (Aβ) peptides encoded by the human transgenes. Additionally, the genetic backgrounds of the different transgenic mice are a possible confounding factor with regard to crucial aspects of AD that they may (or may not) reproduce. Thus, although the usefulness of transgenic mice is undisputed, we hypothesized that additional relevant information on the physiopathology of AD could be obtained from other natural non-transgenic models. We have analyzed the chick embryo and the dog, which may be better experimental models because their enzymatic machinery for processing APP is almost identical to that of humans. The chick embryo is extremely easy to access and manipulate. It could be an advantageous natural model in which to study the cell biology and developmental function of APP and a potential assay system for drugs that regulate APP processing. The dog suffers from an age-related syndrome of cognitive dysfunction that naturally reproduces key aspects of AD including Aβ cortical pathology, neuronal degeneration and learning and memory disabilities. However, dense core neuritic plaques and neurofibrillary tangles have not been consistently demonstrated in the dog. Thus, these species may be natural models with which to study the biology of AD, and could also serve as assay systems for Aβ-targeted drugs or new therapeutic strategies against this devastating disease.

De novo synthesis of GAP-43: in situ hybridization histochemistry and light and electron microscopy immunocytochemical studies in regenerating motor neurons of cranial nerve nuclei in the rat brain.

In order to investigate the modulation of the synthesis and the subcellular localization of the growth associated protein GAP-43 in neuronal cell bodies we have taken advantage of the well known regenerative properties of axotomized motor neurons of the facial and hypoglossal nuclei. Alterations in the levels of GAP-43 mRNA containing cells were studied by in situ hybridization histochemistry. The protein localization was examined using immunohistochemistry at the light and electron microscopic levels. Neurons from the control side showed undetectable levels of both GAP-43-like immunoreactivity and GAP-43 mRNA levels. Whereas axotomized neurons exhibited a marked increase in GAP-43 mRNA levels and in GAP-43-like immunoreactivity. Three to 50 days after axotomy, motor neurons ipsilateral to the lesion displayed a dense reticular or filamentous perinuclear distribution of the immunoreactivity in somata and proximal dendritic processes, corresponding to the location of the Golgi apparatus in these neurons. At the electron microscopic level the immunoreactivity was located in the cisternae of the Golgi complex and found to be associated with trans-side vesicles of these complexes. The myelinated fibers of the transectomized facial nerve also presented an intense GAP-43-like immunoreactivity. Twenty-one days after the axotomy a decay in the number of immunostained neurons and in the intensity of immunolabeled somata was observed. Our study reveals a rapid induction of GAP-43 mRNA and protein after axotomy. The localization of the newly synthesized GAP-43-like immunoreactivity to the Golgi apparatus seen in the present work suggests an early association of this protein with newly formed membranes prior to transport toward the terminals through the axons.

Magnetic resonance spectroscopy in the prediction of early conversion from amnestic mild cognitive impairment to dementia: a prospective cohort study

Background Mild cognitive impairment (MCI) of an amnestic type is a common condition in older people and highly predictive of Alzheimers disease (AD). To date, there is no clear consensus regarding the best antecedent biomarker to predict early conversion to AD. Objective The aim of the study is to demonstrate that 1H magnetic resonance spectroscopy (MRS) of the brain in MCI patients may predict early conversion to dementia within the 2-year period after baseline assessment. Methods A cohort of patients fulfilling the criteria of amnestic MCI were enrolled consecutively. At baseline the patients underwent neuropsychological examination, standard blood tests and APOE genotype. 1H-MRS (1.5 T) of the brain was carried out by exploring two areas: the posteromedial bilateral parietal lobe and left medial occipital lobe. The patients were followed up to detect conversion to probable AD according to the National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimers Disease and Related Disorders Association group criteria. Results After a 2-year follow-up, 27 (38%) patients converted to AD. The mean N-acetyl-aspartate/creatine (NAA/Cr) ratio in the posteromedial bilateral parietal cortex was 1.38 in converters versus 1.49 in non-converters (p<0.0001). An NAA/Cr ratio equal to or lower than 1.43 in this area predicted conversion to probable AD at 74.1% sensitivity and 83.7% specificity (area under the curve: 0.84; 95% CI 0.73 to 0.92). The cross-validated accuracy of classification was 82%, which reaches 85% when the APOE4 genotype and memory test are included in the analysis. In the left medial occipital lobe, the predictive value was somewhat lower with 85.2% sensitivity and 61.4% specificity (area under the curve: 0.8; 95% CI 0.69 to 0.89). Neither the APOE4 genotype nor leuco-araiosis was predictive of conversion to dementia. Conclusion MRS is a valuable biomarker to predict early conversion to dementia in patients with amnestic MCI.

Phosphodiesterase 7 inhibitor reduced cognitive impairment and pathological hallmarks in a mouse model of Alzheimer's disease

Elevated levels of amyloid beta (Aβ) peptide, hyperphosphorylation of tau protein, and inflammation are pathological hallmarks in Alzheimers disease (AD). Phosphodiesterase 7 (PDE7) regulates the inflammatory response through the cyclic adenosine monophosphate signaling cascade, and thus plays a central role in AD. The aim of this study was to evaluate the efficacy of an inhibitor of PDE7, named S14, in a mouse model of AD. We report that APP/Ps1 mice treated daily for 4 weeks with S14 show: (1) significant attenuation in behavioral impairment; (2) decreased brain Aβ deposition; (3) enhanced astrocyte-mediated Aβ degradation; and (4) decreased tau phosphorylation. These effects are mediated via the cyclic adenosine monophosphate/cyclic adenosine monophosphate response element-binding protein signaling pathway, and inactivation of glycogen synthase kinase (GSK)3. Our data support the use of PDE7 inhibitors, and specifically S14, as effective therapeutic agents for the prevention and treatment of AD.

β-amyloid cortical deposits are accompanied by the loss of serotonergic neurons in the dog

Dogs may naturally suffer an age‐related cognitive impairment that has aroused a great deal of interest, even beyond the field of the veterinary clinic. This canine senile dementia reproduces several key aspects of Alzheimers disease (AD), including the presence of β‐amyloid (Aβ) deposits in the cerebral cortex, neurodegeneration, and learning and memory impairments. In the present study, we have used unbiased stereological procedures to estimate the number of the dorsal and median raphe nuclei (DRN and MRN, respectively) serotonergic neurons immunolabeled with an anti‐tryptophan hydroxylase (TrH) monoclonal antibody in young and aged dogs without Aβ cortical deposits and in aged dogs with Aβ cortical deposits. The estimated total number of TrH‐labeled neurons (mean ± SD) was 94,790 ± 26,341 for the DRN and 40,404 ± 8,692 for the MRN. The statistical analyses revealed that aged dogs with Aβ cortical pathology had 33% fewer serotonergic neurons in the DRN and MRN than aged dogs without Aβ cortical deposits (108,043 ± 18,800 vs. 162,242 ± 39,942, respectively; P = 0.01). In contrast, no significant variations were found between young and aged dogs without Aβ cortical deposits. These results suggest that degeneration of the serotonergic neurons could be involved in the cognitive damage that accompanies Aβ cortical pathology in the dog and reinforce the use of the canine model for exploring the potential mechanisms linking the cortical Aβ pathology and serotonergic neurodegeneration that occurs during the course of AD. J. Comp. Neurol. 513:417–429, 2009.

Dogs with canine counterpart of Alzheimer's disease lose noradrenergic neurons.

Degeneration of noradrenergic neurons in the locus ceruleus is a well-described feature of Alzheimers disease (AD). In spite of extensive utilization of the dog as a model for human degenerative diseases, there is no data on the response to aging of the noradrenergic system in dogs. We have used modern unbiased stereology to estimate the total number of A6-A7 noradrenergic neurons in normal, aged dogs and dogs with the canine counterpart of AD. In small-breed dogs with no cognitive impairments, the total mean number of tyrosine hydroxylase immunolabeled A6-A7 neurons was 17,228+/-1655, with no differences between young and aged dogs. In contrast, aged dogs with cognitive impairments exhibited a significant reduction in the total number of A6-A7 neurons (13,487+/-1374; P=0.001). Additionally, we found a negative correlation between the number of A6-A7 neurons and the extent of beta-amyloid deposits in the prefrontal cortex. These results suggest that the canine model could be useful in exploring the potential benefits of noradrenergic drugs for the treatment of AD.

Reliable Measurements of the β-Amyloid Pool in Blood Could Help in the Early Diagnosis of AD

The present study was aimed at assessing the capability of Aβ1-40 and Aβ1-42 levels in undiluted plasma (UP), diluted plasma (DP), and cell bound (CB) to distinguish between early stages of Alzheimers disease (AD), amnesic mild cognitive impairment (MCI), and healthy control (HC). Four blood samples from each participant were collected during one month and the levels of Aβ1-40 and Aβ1-42 were determined by a blinded proprietary ELISA sandwich (Araclon Biotech. Zaragoza, Spain). First striking result was that the amount of Aβ1-40 and Aβ1-42 in UP represented only a small proportion (~15%) of the total beta-amyloid pool in blood (βAPB) described here as the sum of Aβ1-40 and Aβ1-42 in blood where they are free in plasma, bound to plasma proteins, and bound to blood cells. Furthermore, we found that levels of Aβ1-40 and Aβ1-42 in UP, DP, and CB were significantly higher in MCI when compared to HC. On average, the total βAPB was 1.8 times higher in MCI than in HC (P = 0.03) and allowed to discriminate between MCI and HC with a sensitivity and specificity over 80%. Thus, quantification of several markers of the βAPB could be useful and reliable in the discrimination between MCI and HC.

Plasma β-amyloid peptides in canine aging and cognitive dysfunction as a model of Alzheimer's disease

Aging dogs naturally demonstrate cognitive impairment and neuropathology that model early Alzheimers disease (AD). In particular, there is evidence that canine cognitive dysfunction syndrome (CDS) in aged dogs is accompanied by cortical deposition of Aβ peptides and neurodegeneration. Plasma Aβ levels have been examined in humans as putative biomarkers for AD, but to date, no similar studies have been conducted for canine dementia. The aim of the present study was to assess plasma Aβ1-42 and Aβ1-40 levels in a blind study using pet dogs that were either successfully aging or exhibiting CDS. The severity of cognitive impairment was assessed using an owner-based questionnaire. On average, young dogs presented significantly higher plasma levels of Aβ1-42 and Aβ1-40 than aged, cognitively unimpaired dogs. Notably, among aged dogs, the levels of Aβ1-42 and the Aβ42/40 ratio were significantly higher in those showing mild cognitive impairment than in either cognitively unimpaired or severely affected dogs. These results suggest that increased plasma Aβ1-42 levels and Aβ42/40 ratio could be a biomarker for canine cognitive dysfunction, which is considered an excellent natural model of early AD.

Ionotropic glutamate receptor subunit distribution on hypoglossal motoneuronal pools in the rat

The expression of ionotropic glutamate receptor subunits in the motoneuronal pools of the hypoglossal nucleus was studied using specific antibodies against subunits of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), kainate and N-methyl-D-aspartate (NMDA) subtypes. The highest numbers of intensely immunolabelled motoneurons were found in the dorsal tier and caudoventromedial part of the hypoglossal nucleus with all antibodies except that against the GluR1 AMPA subunit. Labelling for the GluR1 subunit was weak except for caudally located groups of motoneurons which innervate tongue muscles related to respiratory activity. By contrast, most motoneurons were intensely immunostained with antibodies against GluR2/3 and GluR4 subunits of the AMPA subtype. The low staining observed using an antibody specific for the GluR2 subunit (which prevents Ca2+-entry through AMPA channels) strongly suggests that AMPA receptors in hypoglossal motoneurons are Ca2+-permeable. Immunolabelling for the GluR5/6/7 kainate receptor subunits was found in many motoneuronal somata as well as in thin axon-like profiles and puncta that resembled synaptic boutons. Most motoneurons were intensely immunostained for the NMDA receptor subunit NR1. These results show that the hypoglossal nucleus contains five heterogeneous pools of motoneurons which innervate functionally defined groups of tongue muscles. The uneven expression of the different receptor subunits analysed here could reflect diverse phenotypic properties of hypoglossal motoneurons which might be expected to generate different patterns of motor responses under different physiological or pathological conditions.