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M Morales

Publications and source records attributed to M Morales.

At least 37 records · Page 2Linked to original sources
PubMed2003

Differential expression of the cell line-derived neurotrophic factor (GDNF) receptor GFRalpha1 in heterozygous Gfralpha1 null-mutant mice after stroke.

Exogenous administration of glial cell line-derived neurotrophic factor (GDNF) reduces ischemia-induced cerebral infarction. Cerebral ischemia induces gene expression of GDNF, GDNF-receptor alpha-1 (GFRalpha-1) and c-Ret, suggesting that a GDNF signaling cascade mechanism may be involved in endogenous neuroprotection during ischemia. In the present study, we examined if this endogenous neuroprotective pathway was altered in Gfralpha-1 deficient mice. Since mice homozygous for the Gfralpha-1 deletion (-/-) die within 24 h of birth, stroke-induced changes in the levels of Gfralpha-1 mRNA were studied in Gfralpha-1 heterozygous (+/-) mice and their wild-type (+/+) littermates. The right middle cerebral artery was transiently ligated for 45 min in anesthetized mice. Animals were killed at 0, 6, 12 and 24 h after the onset of reperfusion and levels of Gfralpha-1 mRNA were measured by in situ hybridization histochemistry. Previously, we showed that Gfralpha-1 (+/-) mice are more vulnerable to focal cerebral ischemia. In the present study, we found that basal levels of GFRalpha-1 mRNA were at similar low levels in cortex and striatum in adult Gfralpha-1 (+/+) and Gfralpha-1 (+/-) mice and that ischemia/reperfusion induced up-regulation of Gfralpha-1 mRNA in the lesioned and contralateral sides of cortex and striatum in both Gfralpha-1 (+/+) and GFRalpha-1 (+/-) mice. However, the ischemia/reperfusion induction of Gfralpha-1 mRNA was significantly higher in the cortex of wild type mice, as compared to Gfralpha-1 (+/-) mice. Moreover, the increased expression of Gfralpha-1 in striatum after reperfusion occurred earlier in the GFRalpha-1 (+/+) than in the Gfralpha-1 (+/-) mice. These results indicate that after ischemia, there is a differential up-regulation of Gfralpha-1 expression in Gfralpha-1 (+/+) and Gfralpha-1 (+/-) mice. Since GDNF has neuroprotective effects, the reduced up-regulation of Gfralpha-1 in Gfralpha-1 (+/-) mice at early time points after ischemia suggests that the responsiveness to GDNF and GDNF receptor mediated neuroprotection is attenuated in these genetically modified animals and may underlie their greater vulnerability.

Animals↗
PubMed2003

HSV amplicon delivery of glial cell line-derived neurotrophic factor is neuroprotective against ischemic injury.

Direct intracerebral administration of glial cell line-derived neurotrophic factor (GDNF) is neuroprotective against ischemia-induced cerebral injury. Utilizing viral vectors to deliver and express therapeutic genes presents an opportunity to produce GDNF within localized regions of an evolving infarct. We investigated whether a herpes simplex virus (HSV) amplicon-based vector encoding GDNF (HSVgdnf) would protect neurons against ischemic injury. In primary cortical cultures HSVgdnf reduced oxidant-induced injury compared to the control vector HSVlac. To test protective effects in vivo, HSVgdnf or HSVlac was injected into the cerebral cortex 4 days prior to, or 3 days, after a 60-min unilateral occlusion of the middle cerebral artery. Control stroke animals developed bradykinesia and motor asymmetry; pretreatment with HSVgdnf significantly reduced such motor deficits. Animals receiving HSVlac or HSVgdnf after the ischemic insult did not exhibit any behavioral improvement. Histological analyses performed 1 month after stroke revealed a reduction in ischemic tissue loss in rats pretreated with HSVgdnf. Similarly, these animals exhibited less immunostaining for glial fibrillary acidic protein and the apoptotic marker caspase-3. Taken together, our data indicate that HSVgdnf pretreatment provides protection against cerebral ischemia and supports the utilization of the HSV amplicon for therapeutic delivery of trophic factors to the CNS.

Animals↗
PubMed2003

Glial cell line neurotrophic factor-family receptor alpha-1 is present in central neurons with distinct phenotypes.

Glial cell line neurotrophic factor(GDNF) is a potent survival factor for several types of neurons. GDNF binds with high affinity to the GDNF-family receptor alpha-1 (GFRalpha-1) which is expressed in different brain areas. In the present study, by using anatomical techniques, we document the phenotypic diversity among GFRalpha-1 expressing neurons in the CNS. GFRalpha-1 expression was found in GABA (gamma-aminobutyric acid)-containing neurons distributed in the cortex, reticular thalamic nucleus and septum. While high expression of GFRalpha-1 was often observed in cholinergic motoneurons in the spinal cord, very few septal cholinergic neurons were found to express GFRalpha-1. GFRalpha-1 transcripts were also detected in catecholaminergic neurons in the periventricular hypothalamic nucleus, dorsal raphe nucleus and locus ceruleus. Within the raphe nucleus, GFRalpha-1 expression was prominent in many serotonergic neurons and in few neurons containing the enzyme nitric oxide synthase. As GFRalpha-1 is activated by GDNF and GDNF-related neurotrophic factors, the widespread distribution of GFRalpha-1 in neurons with different phenotypes indicates that the neuronal activity of these neurons is likely to be affected by GDNF and GDNF-related neurotrophic factors. This would result in the regulation of diverse neuronal pathways in the adult brain. Published by Elsevier Science Ltd on behalf of IBRO.

Acetylcholine↗
PubMed2003

Thyroid hormone modulates ClC-2 chloride channel gene expression in rat renal proximal tubules.

Thyroid hormones has its main role in controlling metabolism, but it can also modulate extracellular fluid Volume (ECFV) through its action on the expression and activity of Na(+) transporters. Otherwise, chloride is the main anion in the ECFV and the influence of thyroid hormones in the regulation of chloride transporters is not yet understood. In this work, we studied the effect of thyroid hormones in the expression of ClC-2, a cell Volume-, pH- and voltage-sensitive Cl(-) channel, in rat kidney. To analyze the modulation of ClC-2 gene expression by thyroid hormones, we used hypothyroid (Hypo) rats with or without thyroxine (T(4)) replacement and hyperthyroid (Hyper) rats as our experimental models. Total RNA was isolated and the expression of ClC-2 mRNA was evaluated by a ribonuclease protection assay, and/or semi-quantitative RT-PCR. Renal ClC-2 expression decreased in Hypo rats and increased in Hyper rats. In addition, semi-quantitative RT-PCR of different nephron segments showed that these changes were due exclusively to the modulation of ClC-2 mRNA expression by thyroid hormone in convoluted and straight proximal tubules. To investigate whether thyroid hormones action was direct or indirect, renal proximal tubule primary culture cells were prepared and subjected to different T(4) concentrations. ClC-2 mRNA expression was increased by T(4) in a dose-dependent fashion, as analyzed by RT-PCR. Western blotting demonstrated that ClC-2 protein expression followed the same profile of mRNA expression.

Animals↗
PubMed2002

Bone morphogenetic proteins are involved in fetal kidney tissue transplantation-induced neuroprotection in stroke rats.

Both bone morphogenetic proteins (BMPs) and glial cell line-derived neurotrophic factor (GDNF) reduce ischemia-induced cerebral injury in rats. Intracerebral transplantation of fetal kidney tissue, which normally expresses BMPs and GDNF during development, reduces ischemic injury in cerebral cortex. In this study, we tested the hypothesis that BMP is involved in this neuroprotective response. Fetal kidney tissue was cut into small pieces and transplanted into cortical areas adjacent to the right middle cerebral artery (MCA) in adult rats. In situ hybridization of brain indicated that these fetal kidney transplants contained high levels of BMP-7 mRNA three days after grafting. Immunohistochemical analysis of grafted brain showed co-localization of BMP-7 and PAX-2 immunoreactivity in the graft, suggesting that these transplants contained BMP protein. Some animals were grafted with fetal kidney tissue after intraventricular administration (ICV) of the BMP antagonist noggin (1 micro g) or after vehicle, followed by MCA ligation for 60 min. Animals receiving fetal kidney tissue transplantation developed significantly less body asymmetry, as compared to stroke animals that either did not receive transplantation or received fetal kidney grafts and noggin pretreatment. Analysis of these brains after triphenyltetrazolium chloride staining showed that fetal kidney tissue transplantation reduced the volume of infarction in the cerebral cortex. Noggin pretreatment reduced the protection induced by fetal kidney grafting, although noggin itself did not cause increase in cerebral infarction. Eight hours after ischemia, brain homogenates were obtained from grafted and control animals to assay caspase-3 enzymatic activity. This analysis demonstrated that fetal kidney grafts significantly reduced ischemia-induced caspase-3 activity. Reduction of caspase-3 activity could also be antagonized by noggin pretreatment. In conclusion, our data suggest that fetal kidney transplantation reduces ischemia/reperfusion-induced cortical infarction and behavioral deficits in adult rats, which are, at least partially, mediated through the effect of BMPs from the transplants.

Animals↗
PubMed2002

Silica urolithiasis in the dromedary camel in a subtropical climate.

In 1998, two cases of silica urolithiasis occurred in castrated male dromedaries on an intensive camel farm in the Canary Islands. The immediate attributable cause was the ingestion of large amounts of silica in the feed, estimated as 84.44 g/day. An associated cause was the low level of salt in the diet. Daily ingestion of salt from feed and water was estimated to be 21.8 g (8.6 g of sodium). Seventy-six castrated males from the same farm were divided into four groups: group A received 30 g of salt daily; group B received 40 g; group C received 60 g; and group D received no added salt in the diet (control). The animals were maintained on these dietary regimes for 2 years. No animals from groups A, B or C suffered overt urinary retention. One animal from group D had an obstructive urinary retention 10 months after the study commenced. Thus, 52 g of salt daily appears to be sufficient to prevent urinary retention in dromedaries raised in a subtropical climate.

Animal Feed↗
PubMed2002

Protective effects of glial cell line-derived neurotrophic factor in ischemic brain injury.

Glial cell line-derived neurotrophic factor (GDNF), a member of the transforming growth factor-beta (TGF-beta) superfamily, has been shown to have trophic activity on dopaminergic neurons. Recent studies indicate that GDNF can protect the cerebral hemispheres from damage induced by middle cerebral arterial ligation. We found that such neuroprotective effects are mediated through specific GDNF receptor alpha-1 (GFRalpha1). Animals with a deficiency in GFRalpha-1 have less GDNF-induced neuroprotection. Ischemia also enhances nitric oxide synthase (NOS) activity, which can be attenuated by GDNF. These.data suggest that GDNF can protect against ischemic injury through a GFRalpha-1/NOS mechanism. We also found that the receptor for GDNF, GFRalpha1, and its signaling moiety c-Ret were upregulated, starting immediately after ischemia. This upregulation suggests that activation of an endogenous neuroprotective mechanism occurs so that responsiveness of GDNF can be enhanced at very early stages during ischemia.

Animals↗
PubMed2002

An atomic force microscopic study of the ultrastructure of dental enamel afflicted with amelogenesis imperfecta.

The ultrastructure of human tooth enamel from a patient diagnosed to have amelogenesis imperfecta (AI) was investigated using atomic force microscopy (AFM) and compared with normal human tooth enamel. AI is a hereditary defect of dental enamel in which the enamel is deficient in either quality or quantity. Tissue-specific proteins, especially amelogenins, have been postulated to play a central role in amelogenesis. The secondary structure of amelogenin has been assigned an important role in directing the architecture of hydroxyapatite (HA) enamel crystallites and an alteration of the secondary structure of amelogenin is expected to result in an altered architecture of the mineral phase in human enamel. Previous studies have shown that the human amelogenin gene encodes for a mutant protein in which a conserved Pro is mutated to a Thr residue (Pro-->Thr); such a mutation should be expected to cause a disoriented pattern of the mineral phase in enamel. AFM results presented for the AI tooth enamel clearly demonstrate that the apatite crystal morphology in AI tooth enamel is perturbed in the diseased state; this might result from a defective synthesis of the extracellular matrix proteins, e.g. amelogenin, by the ameloblasts.

Adult↗
PubMed2002

Glucagon-like peptide-1 (GLP-1) and glucose metabolism in human myocytes.

Glucagon-like peptide-1 (GLP-1) has been shown to have insulin-like effects upon the metabolism of glucose in rat liver, muscle and fat, and on that of lipids in rat and human adipocytes. These actions seem to be exerted through specific receptors which, unlike that of the pancreas, are not - at least in liver and muscle - cAMP-associated. Here we have investigated the effect, its characteristics, and possible second messengers of GLP-1 on the glucose metabolism of human skeletal muscle, in tissue strips and primary cultured myocytes. In muscle strips, GLP-1, like insulin, stimulated glycogen synthesis, glycogen synthase a activity, and glucose oxidation and utilization, and inhibited glycogen phosphorylase a activity, all of this at physiological concentrations of the peptide. In cultured myotubes, GLP-1 exerted, from 10(-13) mol/l, a dose-related increase of the D-[U-(14)C]glucose incorporation into glycogen, with the same potency as insulin, together with an activation of glycogen synthase a; the effect of 10(-11) mol/l GLP-1 on both parameters was additive to that induced by the equimolar amount of insulin. Synthase a was still activated in cells after 2 days of exposure to GLP-1, as compared with myotubes maintained in the absence of peptide. In human muscle cells, exendin-4 and its truncated form 9-39 amide (Ex-9) are both agonists of the GLP-1 effect on glycogen synthesis and synthase a activity; but while neither GLP-1 nor exendin-4 affected the cellular cAMP content after 5-min incubation in the absence of 3-isobutyl-1-methylxantine (IBMX), an increase was detected with Ex-9. GLP-1, exendin-4, Ex-9 and insulin all induced the prompt hydrolysis of glycosylphosphatidylinositols (GPIs). This work shows a potent stimulatory effect of GLP-1 on the glucose metabolism of human skeletal muscle, and supports the long-term therapeutic value of the peptide. Further evidence for a GLP-1 receptor in this tissue, different from that of the pancreas, is also illustrated, suggesting a role for an inositolphosphoglycan (IPG) as at least one of the possible second messengers of the GLP-1 action in human muscle.

Analysis of Variance↗
PubMed2001

GFRalpha-1 mRNA in dopaminergic and nondopaminergic neurons in the substantia nigra and ventral tegmental area.

Glial cell line-derived neurotrophic factor (GDNF) is a survival factor for several types of neurons, including dopaminergic (DAergic) neurons. GDNF binds with high affinity to the GDNF family receptor alpha-1 (GFRalpha-1), which is highly expressed in the midbrain. Using anatomical and lesion techniques, we demonstrated that GFRalpha-1 was expressed in DAergic and non-DAergic neurons in the rat midbrain. Immunohistochemical characterization of GFRalpha-1-expressing neurons indicated that most of the neurons that were immunopositive for the DAergic marker tyrosine hydroxylase (TH) expressed GFRalpha-1 in the substantia nigra pars compacta (SNC). In contrast, fewer TH-containing neurons expressed GFRalpha-1 in the substantia nigra pars reticulata (SNR) and the ventral tegmental area (VTA). Depletion of GFRalpha-1/TH neurons was observed in the SNC following treatment with the neurotoxin 6-hydroxydopamine (6-OHDA); however, GFRalpha-1 expression remained in some neurons located in the SNR. The gamma-aminobutyric acid (GABA)ergic nature of GFRalpha-1-expressing neurons located in the SNR, which were resistant to (6-hydroxydopamine) 6-OHDA, was established by their expression of glutamic acid decarboxylase (GAD; the synthesizing enzyme for GABA). Further analysis indicated that coexpression of GFRalpha-1 and GAD varied in a rostrocaudal gradient in the SNR, substantia nigra pars lateralis (SNL), and VTA. Midbrain DAergic and GABAergic neurons have been previously classified according to their Ca(2+) binding protein (CaBP) content; thus, we also sought to investigate the proportion of midbrain GFRalpha-1-expressing neurons containing parvalbumin (PV), calbindin (CB), and calretinin (CR) in the midbrain. Although GFRalpha-1 expression was found mainly in CB- and CR-immunoreactive neurons, it was rarely observed in PV-immunolabeled neurons. Analysis of the proportion of GFRalpha-1-expressing neurons for each CaBP subpopulation indicated the coexistence of GFRalpha-1 with CR in the VTA and all subdivisions of the SN; double-labeled GFRalpha-1/CR neurons were distributed in the SNC, SNR, SNL, and VTA. GFRalpha-1/CB neurons were also detected in the SNC, SNL, and VTA. Expression of GFRalpha-1 in DAergic and non-DAergic neurons in the rat SN and VTA suggests that GDNF, via GFRalpha-1, might modulate DAergic and GABAergic functions in the nigrostriatal, mesolimbic, and nigrothalamic circuits of the adult rat.

Animals↗
PubMed2001

Histochemical differentiation of Dirofilaria immitis, Dirofilaria repens and Acanthocheilonema dracunculoides microfilariae by staining with a commercial kit, Leucognost-SP.

The diagnosis of canine heartworm infection is based upon the presence of circulating Dirofilaria immitis microfilariae or on techniques for the detection of serum antibodies or antigens. In the first of these, discrimination between D. immitis, D. repens and Acanthocheilonema dracunculoides microfilariae is based upon the acid phosphatase histochemical stain. In this paper, we propose an alternative technique for histochemical staining using a commercial kit test of naphthol-AS-OL (Leucognost-SP). This offers the advantages of speed and simplicity as compared to the standard Barka procedure.

Acid Phosphatase↗
PubMed2001

Interactions between dopamine D1 receptors and gamma-aminobutyric acid mechanisms in substantia nigra pars reticulata of the rat: neurochemical and behavioral studies.

RATIONALE: Several studies have shown that dopamine D1 agonists act on forebrain dopamine terminal regions to exert many of their behavioral effects. Yet, there is also a large number of D1 receptors in the substantia nigra pars reticulata (SNr), and these receptors are located mainly on terminals of gamma-aminobutyric acid (GABA)-ergic striatonigral neurons. OBJECTIVE: The present studies were undertaken to determine the behavioral and neurochemical effects of local administration of the D1 agonist SKF 82958 and to study the interactions between D1 and GABA mechanisms in SNr. METHODS: Microdialysis methods were used to characterize the effect of SKF 82958 on extracellular GABA, and several experiments studied the effects of nigral D1 stimulation on motor activity and investigated the behavioral significance of D1/GABA interactions in SNr. RESULTS: Local infusion of 10(-6) M SKF 82958 increased extracellular levels of SNr GABA, and this effect was blocked by co-infusion of the D1 antagonist SCH 23390. Bilateral SNr injections of SKF 82958 increased locomotor activity, and this effect was blocked by the GABA-A antagonist bicuculline. Intranigral bicuculline reduced motor activity, while the GABA-A agonist muscimol increased various motor activities in a manner similar to SKF 82958. CONCLUSIONS: The present results suggest that the D1 agonist SKF 82958 acts on D1 receptors in SNr to increase extracellular levels of GABA, and the increase in motor activity produced by nigral D1 stimulation is dependent on stimulation of GABA-A receptors. D1/GABA interactions in SNr are important for the modulation of basal ganglia output, which may have important implications for Parkinson's disease.

Animals↗
PubMed2001

5-HT(3)-receptor subunits A and B are co-expressed in neurons of the dorsal root ganglion.

The type 3 serotonin (5-HT(3)) receptor is the only ligand-gated ion channel receptor for serotonin (5-HT). Many pharmacological, behavioral, and electrophysiological studies indicate heterogeneous properties for this receptor. Although the basis for this heterogeneity is unknown, one possible explanation for these findings resides in the subunit composition of the receptor. Two 5-HT(3)-receptor subunits have been cloned: the 5-HT(3)-receptor subunit A (5-HT(3A)) and the 5-HT(3)-receptor subunit B (5-HT(3B)). Recombinant co-expression of 5-HT(3A) and 5-HT(3B) subunits produces a functional heteromeric 5-HT(3A/3B) receptor with pharmacological and electrophysiological properties different from those displayed by the 5-HT(3A) homomeric receptor. In the present report, we used in situ hybridization histochemistry to demonstrate that the 5-HT(3B) subunit is expressed in rat dorsal root ganglion (DRG) neurons. We determined with cellular resolution that 5-HT(3B) subunit mRNA was expressed in 43.2 +/- 2.8% of the total population of DRG neurons. By comparison, the 5-HT(3A) subunit was more widely expressed, with 70.0 +/- 2.8% of the total population of DRG neurons expressing this subunit. Further analyses showed that most of the neurons containing mRNA for the 5-HT(3B) subunit (91.5 +/- 3.4%) also expressed the 5-HT(3A) subunit. In contrast, nearly half the population of neurons expressing 5-HT(3A) subunit lacked (52.8 +/- 5.9%) transcripts for the 5-HT(3B) subunit. These results provide the first evidence indicating that the 5-HT(3B) subunit of the 5-HT(3) receptor is expressed in DRG and suggest that sensory neurons have the capacity to synthesize at least two structurally different 5-HT(3) receptors: a heteromeric 5-HT(3A/3B) receptor and a homomeric 5-HT(3A) receptor. Consequently, 5-HT(3) receptors with different properties might be present in peripheral and central axons of the DRG. These findings open the possibility that distinct types of 5-HT(3) receptors may be involved in perception and/or processing of sensory information. J. Comp. Neurol. 438:163-172, 2001. Published 2001 Wiley-Liss, Inc.

Animals↗
PubMed2001

First field trial of a transmissible recombinant vaccine against myxomatosis and rabbit hemorrhagic disease.

As a novel approach for immunisation of wild rabbits, we have recently developed a transmissible vaccine against myxomatosis and rabbit hemorrhagic disease (RHD) based on a recombinant myxoma virus (MV) expressing the RHDV capsid protein [J. Virol. 74 (2000) 1114]. The efficacy and safety of the vaccine have been extensively evaluated under laboratory conditions. In this study, we report the first limited field trial of the candidate vaccine that was undertaken in an island of 34 Has containing a population of around 300 rabbits. Following administration by the subcutaneous route to 76 rabbits, the vaccine induced specific antibody responses against both myxomatosis and RHDV in all the inoculated rabbits. Furthermore, the recombinant virus exhibited a limited horizontal transmission capacity, promoting seroconversion of around 50% of the uninoculated rabbit population. No evidence of undesirable effects due to the recombinant virus field release was detected.

Animals↗
PubMed2001

Cellular mRNA expression of the transcription factor NGFI-B suggests a gene regulatory role in striatal opiate-peptide neurons.

Previous studies have shown that NGFI-B mRNA is highly expressed in the adult striatum. In the present study we analyzed the anatomical distribution of NGFI-B mRNA within this brain region as well as the degree of co-existence of NGFI-B with different striatal markers in the adult brain. NGFI-B mRNA levels were found to be significantly higher within the dorsomedial portion of the striatum as compared to the ventrolateral striatum. This distribution pattern was maintained throughout the rostro--caudal axis of the striatum. Double in situ hybridization studies showed that striatal NGFI-B mRNA colocalized with a subset of preproenkephalin and prodynorphin positive spiny neurons within the dorsomedial striatum; 22--28% of all opiate-peptide positive cells co-expressed NGFI-B mRNA. NGFI-B did not colocalize with striatal aspiny interneurons expressing choline acetyl transferase mRNA or those containing the calcium-binding protein parvalbumin. The pattern of NGFI-B mRNA expression within different striatal spiny projecting neurons suggests that this transcription factor may have a direct effect on the function of different striatal efferent pathways.

Animals↗