Biomedical subjects

F J Alvarez

Publications and source records attributed to F J Alvarez.

At least 19 recordsLinked to original sources

PubMed2000

Factors regulating AMPA-type glutamate receptor subunit changes induced by sciatic nerve injury in rats.

Excitatory glutamatergic neurotransmission at Ia afferent-motoneuron synapses is enhanced shortly after physically severing or blocking impulse propagation of the afferent and/or motoneuron axons. We considered the possibility that these synaptic changes occur because of alterations in the number or properties of motoneuron alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) receptors. Therefore, we quantitatively analyzed glutamate receptor (GluR)1, GluR2/3, and GluR4 AMPA subunit immunoreactivity (ir) in motoneurons 3, 7, or 14 days after axotomy or continuous tetrodotoxin (TTX) block of the sciatic nerve. GluR1-ir remained low in experimental and control motoneurons with either treatment and at any date. However, there was a large reduction of GluR2/3-ir (peak at 7 days >60% reduced) and a smaller, but statistically significant, reduction of GluR4-ir (around 10% reduction at days 3, 7, and 14) in axotomized motoneurons. TTX sciatic blockade did not affect AMPA subunit immunostainings. Axonal injury or interruption of the trophic interaction between muscle and spinal cord, but not activity disruption, appears therefore more likely responsible for altering AMPA subunit immunoreactivity in motoneurons. These findings also suggest that synaptic plasticity induced by axotomy or TTX block, although similar in the first week, could be related to different mechanisms. The effects of axotomy or TTX block on motoneuron expression of the metabotropic glutamate receptor mGluR1a were also studied. mGluR1a-ir was also strongly decreased after axotomy but not after TTX treatment. The time course of the known stripping of synapses from the cell somas of axotomized motoneurons was studied by using synaptophysin antibodies and compared with AMPA and mGluR1a receptor changes. Coverage by synaptophysin-ir boutons was only clearly decreased 14 days post axotomy and not at shorter intervals or after TTX block.

Animals↗

PubMed2000

Postnatal maturation of gephyrin/glycine receptor clusters on developing Renshaw cells.

Adult mammalian Renshaw cells express large and complex postsynaptic gephyrin/glycine receptor clusters on their surface. Larger gephyrin clusters correlate with more "efficacious" inhibitory synapses, in terms of larger postsynaptic quantal size amplitudes, in part because they likely contain more postsynaptic receptors (Lim et al. [1999] J. Physiol. (Lond.) 516:505-512; Oleskevich et al. [1999] J. Neurophysiology 82:312-319). Here, we studied the postnatal development of the gephyrin/glycine receptor cluster size on Renshaw cells. Renshaw cells were identified by their calbindin immunoreactivity, location and morphology, and presence of cholinergic input. The populations of clusters over developing Renshaw cells immunoreactive to gephyrin or glycine receptor alpha1 subunits were comparable in number, size, and complexity and displayed a high degree of colocalization (>90%) at all ages. Quantitative morphologic analysis was performed on gephyrin-immunoreactive clusters. In neonatal animals, Renshaw cells expressed small punctate gephyrin-immunoreactive clusters (mean cluster size +/- SD = 0.19 +/- 0.19 microm(2)at 2 days; 0.22 +/- 0. 19 microm(2)at 5 days). By 10 and 15 days of age, Renshaw cells exhibited gephyrin-immunoreactive clusters that were larger and more complex (0.32 +/- 0.19 microm(2) at 10 days; 0.41 +/- 0.32 microm(2) at 15 days). Cluster growth reached a plateau in 25- and 60-day-old Renshaw cells (0.45 +/- 0.43 microm(2); 0.56 +/- 0.55 microm(2), respectively). By using electron microscopy, we confirmed that gephyrin-immunoreactive clusters were located at postsynaptic sites at both early and late postnatal ages on Renshaw cells. The potential significance of this gephyrin/glycine receptor cluster size maturation that sets Renshaw cells apart from other interneurons is discussed.

Animals↗

PubMed2000

Differential distribution of metabotropic glutamate receptors 1a, 1b, and 5 in the rat spinal cord.

Metabotropic glutamate receptors (mGluRs) modulate somatosensory, autonomic, and motor functions at spinal levels. mGluR postsynaptic actions over spinal neurons display the pharmacologic characteristics of type I mGluRs; however, the spinal distribution of type I mGluR isoforms remains poorly defined. In this study, the authors describe a differential distribution of immunoreactivity to various type I mGluR isoforms (mGluR1a, mGluR5a,b, and mGluR1b) that suggests a correlation between specific isoforms and particular aspects of spinal cord function. Two different antisera raised against mGluR5a,b detected intense immunoreactivity within nociceptive afferent terminal fields (laminae I and II) and also in autonomic regions (parasympathetic and sympathetic). In contrast, two of three anti-mGluR1a antibodies did not immunostain lamina I or II. Laminae I and II immunostaining by a third anti-mGluR1a antibody was competed by a peptide sequence obtained from a homologous region in mGluR5, suggesting possible cross reactivity in fixed tissue. Autonomic neurons did not express mGluR1a immunoreactivity. All anti-mGluR1a antibodies strongly and specifically immunolabeled dendritic and somatic membranes of neurons in the deep dorsal horn (lamina III-V) and the ventral horn (lamina VI-IX). Somatic motoneurons expressed mGluR1a immunoreactivity but little or no mGluR5 immunoreactivity. Phrenic and pudendal motoneurons expressed the highest level of mGluR1a immunoreactivity in the spinal cord. Intense mGluR1b immunoreactivity was restricted to a few scattered neurons and a prominent group of neurons in lamina X. Lamina II neurons expressed low levels of mGluR1b immunoreactivity. Ultrastructurally, type I mGluR immunoreactivity was found mostly at extrasynaptic sites on the plasma membrane, but it was also found perisynaptically, in the body of the postsynaptic regions or in relation to intracytoplasmic structures.

Animals↗

PubMed2000

GABA mediates presynaptic inhibition at glycinergic synapses in a rat auditory brainstem nucleus.

Many inhibitory nerve terminals in the mammalian anteroventral cochlear nucleus (AVCN) contain both glycine and GABA, but the reason for the co-localization of these two inhibitory neurotransmitters in the AVCN is unknown. We have investigated the roles of glycine and GABA at synapses on bushy cells in the rat AVCN, using receptor immunohistochemistry and electrophysiology. Our immunohistochemical results show prominent punctate labelling of postsynaptic clusters of glycine receptors and of the receptor clustering protein gephyrin over the surface of bushy cells. In contrast, weak diffuse membrane immunolabelling of GABAA receptors was observed. Whole-cell recordings from bushy cells in AVCN slices demonstrated that evoked inhibitory postsynaptic currents (IPSCs) were predominantly (81 %) glycinergic, based on the decrease in amplitude of the IPSCs in bicuculline (10 microM). This observation was supported by the effect of strychnine (1 microM), which was to decrease the evoked IPSC (to 10 % of control IPSC amplitude) and to produce a greater than 90 % block of spontaneous miniature IPSCs. These results suggest a minor role for postsynaptic GABAA receptors in bushy cells, despite a high proportion of GABA-containing terminals on these cells. Therefore, a role for metabotropic GABAB receptors was investigated. Activation of GABAB receptors with baclofen revealed a significant attenuation of evoked glycinergic IPSCs. The effect of baclofen was presynaptic, as indicated by a lack of change in the mean amplitude of spontaneous IPSCs. Significantly, the decrease in the amplitude of evoked glycinergic IPSCs observed following repetitive nerve stimulation was reduced in the presence of the GABAB antagonist, CGP 35348. This indicates that synaptically released GABA can activate presynaptic GABAB receptors to reduce transmitter release at glycinergic synapses. Our results suggest specific pre- versus postsynaptic physiological roles for GABA and glycine in the AVCN.

Animals↗

PubMed2000

Presence of illegal drugs in drivers involved in fatal road traffic accidents in Spain.

This study investigated the presence of illegal drugs in the blood of 285 fatally injured drivers in Spain. Illegal drugs were detected in 10.2% of all samples. Illicit drugs alone were detected in 2.5% and together with other substances in 7.7%. Cocaine was the most common drug detected. The mean number (+/- S.D.) of substances detected was 2.6 +/- 1.2: consisting of 46 illegal drugs, 14 alcohol cases and 16 medicines. Three concentration levels of the different substances have been established: low, medium and high-toxic. In 68.9% of the samples in which an illegal drug was detected, a substance was also found at the high-toxic level. The results show that illegal drugs are commonly detected in road accident victims.

Accidents, Traffic↗

PubMed2000

Ultrastructural localization of monoclonal antiphospholipid antibody binding to rat brain.

Antiphospholipid antibodies (aPL), in the presence or absence of systemic lupus erythematosus, are associated with a number of neurologic complications. However, the role aPL play in pathology is unclear. A thrombotic etiology seems likely for many associated disorders, but not for others. Here we describe aPL-reactive sites in the central nervous system (CNS). Previously, using light microscopy, we showed direct binding of two monoclonal phosphatidylserine-reactive antibodies (aPS) to ependyma and myelin of fixed cat brain. In this study we determined the ultrastructural localization of their binding sites in rat CNS using immunogold electron microscopy techniques. Both monoclonal antibodies reacted strongly with myelin, preferentially with the major dense line formed by the cytoplasmic apposition of the oligodendrocyte plasma membrane. Both monoclonal antibodies also reacted with an antigen that appears associated with the axoneme in cilia of ependymal and choroid plexus epithelium. One monoclonal aPS also showed some reactivity with brain vascular endothelium and reacted slightly with mitochondria, while the other aPS did not react with these structures. While the etiology of aPL-associated neurologic disorders remains unclear, our data suggest possible target sites within the CNS with which aPL can react.

Animals↗

PubMed2000

Nociceptors for the 21st century.

This review summarizes recent developments in the context of the neurochemical classification of nociceptors and explores the relationships between functionally and neurochemically defined subgroups. Although the complete picture is not yet available, several lines of intriguing evidence suggest that despite the complexity and diversity of nociceptor properties, a relatively "simple" neurochemical classification fits well with several recently identified molecular characteristics.

Animals↗

PubMed2000

Influence of aerobic oxidation of mouse erythrocytes on their recognition by macrophages.

Membrane protein modification can change cell surface properties which can be correlated with altered macrophage-erythrocyte interactions. Mouse erythrocytes were incubated in phosphate buffer for different times to induce protein modification. Mouse erythrocyte membrane changes were analyzed by infrared analyses and gel electrophoresis. Proteolytic digestion of membrane proteins was observed. After 22 hours preliminary incubation, the number of erythrocytes adhering to a monolayer of macrophages reached a maximum, the majority of which had not been phagocytosed. Most of the erythrocytes incubated for 40 hours underwent phagocytosis after adhesion to the macrophages.

Animals↗

PubMed1999

Alcohol use among fatally injured drivers in Spain.

Blood from 285 fatally injured drivers in Northern Spain was collected and tested for the presence of alcohol and drugs. Alcohol was detected in 50.5% of all fatalities. Alcohol alone was detected in 44.2% of all samples and in the remaining 6.3% another substance was found together with alcohol. Blood alcohol concentration was classified in different levels. It has been observed that in 35.4% of the cases the blood alcohol level was > or = 0.8 g/l, the legal limit in Spain for car drivers. Alcohol together with other substances was encountered in 18 cases, with medication in 22.2% (4 out of 18), alcohol with illegal drugs in 66.6% of the cases (12 out of 18), and alcohol with medicines and illegal drugs in 11.1% (2 out of 18). Cocaine was the most commonly detected drug. The study shows how widespread the incidence of a high level of alcohol concentration among drivers involved in fatal accidents in Spain.

Accidents, Traffic↗

PubMed1999

Quantal size is correlated with receptor cluster area at glycinergic synapses in the rat brainstem.

1. Whole-cell patch electrode recordings of glycinergic miniature inhibitory postsynaptic currents (mIPSCs) were obtained in neurons of the rat anteroventral cochlear nucleus (AVCN). Mean mIPSC peak amplitude was found to vary considerably between AVCN neurons (range, -19.1 to -317.9 pA; mean +/- s.d., -159.1 +/- 100.7 pA; 14 cells). 2. Immunolabelling of glycinergic receptor clusters in AVCN neurons was performed using antibodies against the glycine receptor clustering protein gephyrin. Measurements of the area of gephyrin immunoreactive clusters were obtained using confocal fluorescence microscopy. These measurements showed a large variability in cluster area, not only in the same cell (mean coefficient of variation, c.v., 0.66 +/- 0.18; 16 cells), but also in mean cluster area between cells (range, 0.21-0.84 microm2; 16 cells). 3. A possible relationship between mIPSC amplitude and receptor cluster area was investigated in a further series of experiments, in which mIPSCs recordings and immunolabelling of glycine receptor clusters were obtained for the same cells. In these experiments, AVCN neurons were identified using intracellular labelling with neurobiotin. Successful results using a combination of whole-cell recordings, neurobiotin identification and immunolabelling were obtained for a total of 10 AVCN neurons. Analysis of the results revealed a positive, statistically significant correlation between mean receptor cluster size and mean mIPSC amplitude (P < 0.05, 10 cells, Spearman's correlation test). 4. These results provide direct experimental evidence supporting a hypothesis of central glycinergic transmission in which synaptic strength may be regulated by changes in the size of the postsynaptic receptor region.

Animals↗

PubMed1999

Distribution of cholinergic contacts on Renshaw cells in the rat spinal cord: a light microscopic study.

1. Cholinergic terminals in the rat spinal cord were revealed by immunohistochemical detection of the vesicular acetycholine transporter (VAChT). In order to determine the relationships of these terminals to Renshaw cells, we used dual immunolabelling with antibodies against gephyrin or calbindin D28k to provide immunohistochemical identification of Renshaw cells in lamina VII of the ventral horn. 2. A total of 50 Renshaw cells were analysed quantitatively using a computer-aided reconstruction system to provide accurate localization of contact sites and determination of somatic and dendritic surface area. Dendrites could be traced for up to 413 microm from the soma in calbindin D28k-identified Renshaw cells and up to 184 microm in gephyrin-identified cells. 3. A total of 3330 cholinergic terminals were observed on 50 Renshaw cells, with a range of 21-138 terminal appositions per cell (mean 66.6 +/- 25.56 contacts per cell). The vast majority (83.5 %) of the terminals were apposed to dendrites rather than the soma. The overall density of cholinergic contacts increased from a little above 1 per 100 microm2 on the soma and initial 25 microm of proximal dendrites to 4-5 per 100 microm2 on the surface of dendritic segments located 50-250 microm from the soma. Single presynaptic fibres frequently formed multiple contacts with the soma and/or dendrites of individual Renshaw cells. 4. VAChT-immunoreactive terminals apposed to Renshaw cells varied in size from 0.6 to 6.9 microm in diameter (mean 2.26 +/- 0.94; n = 986) and were on average smaller than the cholinergic C-terminals apposed to motoneurones, but larger than VAChT-immunoreactive terminals contacting other ventral horn interneurones. 5. The high density and relatively large size of many cholinergic terminals on Renshaw cells presumably correlates with the strong synaptic connection between motoneurones and Renshaw cells. The fact that the majority of contacts are distributed over the dendrites makes the motoneurone axon collateral input susceptible to inhibition by the prominent glycinergic inhibitory synapses located on the soma and proximal dendrites. The relative positions and structural features of the excitatory cholinergic and inhibitory glycinergic synapses may explain why Renshaw cells, although capable of firing at very high frequency following motor axon stimulation, appear to fire at relatively low rates during locomotor activity.

Animals↗

PubMed1999

Diagnosis of attention-deficit/hyperactivity disorder and use of psychotropic medication in very young children.

CONTEXT: Increases in diagnosis and treatment of attention-deficit/hyperactivity disorder (ADHD) have elicited public and professional concern. Research suggests that this trend warrants the inclusion of previously underdiagnosed children and adults. It is not clear whether this trend includes young children. OBJECTIVE: To identify patterns of diagnosis and treatment of ADHD in very young children over time. DESIGN: Descriptive study of Michigan Medicaid claims data. PATIENTS: Inclusion criteria included recorded ADHD diagnosis, continuous Medicaid eligibility during a 15-month period, and age 3 years or younger at the first date of service. MAIN OUTCOME MEASURES: Diagnoses of ADHD, conditions commonly comorbid with ADHD, other chronic health conditions, and injuries; treatments such as psychological services and psychotropic medication; and the number of ambulatory visits. RESULTS: We identified 223 children aged 3 years or younger diagnosed with ADHD. Many had conditions commonly comorbid with ADHD (44%), other chronic health conditions (41%), and injuries (40%). More than half received psychotropic medication (57%); fewer received psychological services (27%). Twenty-two different psychotropic medications were used. Patterns included more than 1 psychotropic medication (46%) in 30 combinations of simultaneous use and 44 combinations of sequential use. The mean number of ambulatory visits was 18. CONCLUSIONS: Children aged 3 years or younger had ADHD diagnosed and received markedly variable psychotropic medication regimens. Little information is available to guide these practices. The presence of comorbid conditions and injuries attests to these children's vulnerability. Resources must be identified that will enable physicians to better respond to the compelling needs of these children and their families.

Age Factors↗

PubMed1999

Allergy of anaesthetizing agents in Spain.

We have investigated the incidence of requests for allergy testing in 5005 patients attending an anaesthetic assessment clinic. Diagnosis of allergy to anaesthetic drugs was established using cutaneous tests. Allergy tests were requested in 151 (3.0%) patients, proving positive in 43 (0.86%). No allergic reactions were observed during subsequent anaesthesia.

Adolescent↗

PubMed1999

Glycinergic miniature synaptic currents and receptor cluster sizes differ between spinal cord interneurons.

The structural features of a synaptic connection between central neurons play an important role in determining the strength of the connection. In the present study, we have examined the relationship between the structural and functional properties of glycinergic synapses in the rat spinal cord. We have analyzed the structure of glycinergic receptor clusters on rat ventral horn interneurons using antibodies against the glycine receptor clustering protein, gephyrin. We have examined the properties of quantal glycinergic currents generated at these synapses using whole cell patch-clamp recordings of miniature postsynaptic inhibitory currents (mIPSCs) in rat spinal cord slices in vitro. Our immunolabeling results demonstrate that there is a considerable variability in the size of glycine receptor clusters within individual neurons. Furthermore there are large differences in the mean cluster size between neurons. These observations are paralleled closely by recordings of glycinergic mIPSCs. The mIPSC amplitude varies significantly within and between neurons. Results obtained using combined immunolabeling and electrophysiological recording on the same neurons show that cells with small glycine receptor clusters concurrently exhibit small mIPSCs. Our results suggest that the differences in the size of glycinergic receptor clusters may constitute an important factor contributing to the observed differences in mIPSC amplitude among spinal cord interneurons.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1999

Substance use by Spanish university students.

The major studies investigating substance use among Spanish university students are reviewed, their results and limitations noted, and current and future substance use trends are discussed.

Behavior, Addictive↗

PubMed1999

Illegal drug-using trends among students in a Spanish university in the last decade (1984-1994).

A sample of 2,086 university students in Valladolid (Spain) were surveyed in 1994 to assess their current use of illicit drugs. That information was used as a baseline to show the trends in the last decade in order to compare two other studies carried out on a similar target population in 1984 and 1990. Of those surveyed, 28.3% had taken some illicit drug within their lifetime, 16.7% in the previous year, and 7.2% in the previous month. Cannabis was the most common illicit drug used in the three levels among these students. 14.2 is the average starting age at which inhalants are used and 19.3 for opiates. 49.8% were opposed to any drug legalization. More than a quarter of the students (28.7%) could be considered as a mental disorder case-finding as measured by Golberg's General Health Questionnaire (GHQ), which was much more relevant among illicit drug users than among nonusers. A decrease in illegal drug use frequency among university students has been observed in the last 10 years.

Adolescent↗

PubMed1999

Band-3 crosslinking-induced targeting of mouse carrier erythrocytes.

Mouse band-3 crosslinked carrier erythrocytes have been prepared. [125I]Carbonic anhydrase (CA) has been encapsulated into mouse erythrocytes. Then, loaded erythrocytes were labelled with 51Cr. Eventually, these doubly labelled cells were crosslinked with band-3 crosslinking reagents. [125I]CA was shown to have cytosolic localization in crosslinked carrier erythrocytes. Estimation of the action of band-3 crosslinkers on mouse carrier-erythrocyte membranes rendered values around 1721% of band-3 monomer reduction. Crosslinked carrier erythrocytes were in vivo targeted to liver, as shown by chromium-labelling localization. Also, encapsulated CA radioactivity was localized in vivo predominantly in liver, which is clearly in contrast with the behaviour shown by free CA injected into animals. These results support this model as a feasible system for the analysis of carrier-erythrocyte survival and targeting as well as the in vivo efficacy of release and targeting of encapsulated compounds.

Animals↗

PubMed1998

Effects of botulinum neurotoxin type A on the expression of gephyrin in cat abducens motoneurons.

In this study, we investigated the effects of long-term synaptic blockade on postsynaptic receptor clustering at central inhibitory glycinergic synapses. High doses of botulinum neurotoxin type A injected in the lateral rectus muscle completely abolishes inhibitory postsynaptic potentials onto abducens motoneurons within 2 days postinjection, and transmission remains blocked for at least 2 months. Using this model, we analyzed the expression of gephyrin, a glycine receptor clustering protein, on the membrane of motoneuron somata after botulinum neurotoxin type A injection in their target muscle. Immunofluorescence or electron microscopy immunohistochemistry revealed gephyrin-immunoreactive clusters (most < 0.5 microm in diameter) densely covering the surface of control abducens motoneurons. Ultrastructurally, presynaptic terminals containing flattened synaptic vesicles (F terminals) were found associated with multiple gephyrin-immunoreactive postsynaptic densities (average 1.24 gephyrin clusters/F+ profile). No significant changes in gephyrin-immunoreactive clusters were observed at 5 days postinjection, but we found significant reductions (25-40%) in the density of gephyrin clusters 19 and 35 days postinjection. Hence, the physiological alterations reported in this model precede structural changes on postsynaptic receptor cluster density. The decrease in gephyrin-immunoreactive clusters was paralleled by reductions in synaptic covering (F+ terminals per 100 microm of membrane). Presumed inactive F+ terminals that remained attached to the motoneuron surface displayed normal gephyrin-immunoreactive clusters; however, the pre- and postsynaptic membranes in between synaptic active zones frequently appeared separated by enlarged extracellular spaces. We concluded that postsynaptic receptor cluster dissolution seemed more directly related to terminal retraction than to inactivity alone.

Abducens Nerve↗