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Biomedical subjects

A M Craig

Publications and source records attributed to A M Craig.

At least 55 records · Page 3Linked to original sources
PubMed1998

Postsynaptically silent synapses in single neuron cultures.

We have used the synapses that isolated hippocampal cells in culture form onto themselves (autapses) to determine if some synapses lack functional AMPA receptors (AMPARs). A comparison of the synaptic variability of the AMPAR- and NMDAR-mediated evoked responses, as well as of miniature synaptic responses, indicates that a population of events exists that only contains an NMDAR component. Spillover of glutamate from adjacent synapses cannot explain these results because in single cell cultures all synaptic events mediated by AMPARs should be detected. Immunocytochemical analysis of these cultures clearly reveals a population of synapses with puncta for NR1 (NMDAR) but not for GluR1 (AMPAR). These results provide strong anatomical and physiological evidence for the existence of postsynaptically silent synapses.

Animals↗
PubMed1998

CRIPT, a novel postsynaptic protein that binds to the third PDZ domain of PSD-95/SAP90.

The synaptic protein PSD-95/SAP90 binds to and clusters a variety of membrane proteins via its two N-terminal PDZ domains. We report a novel protein, CRIPT, which is highly conserved from mammals to plants and binds selectively to the third PDZ domain (PDZ3) of PSD-95 via its C terminus. While conforming to the consensus PDZ-binding C-terminal sequence (X-S/T-X-V-COOH), residues at the -1 position and upstream of the last four amino acids of CRIPT determine its specificity for PDZ3. In heterologous cells, CRIPT causes a redistribution of PSD-95 to microtubules. In brain, CRIPT colocalizes with PSD-95 in the postsynaptic density and can be coimmunoprecipitated with PSD-95 and tubulin. These findings suggest that CRIPT may regulate PSD-95 interaction with a tubulin-based cytoskeleton in excitatory synapses.

Adaptor Proteins, Signal Transducing↗
PubMed1998

Blood mineral and vitamin E concentrations in llamas.

OBJECTIVE: To establish reference values for blood concentrations of total calcium (Ca), inorganic phosphorus (P), iron (Fe), copper (Cu), zinc (Zn), selenium (Se), and vitamin E (Vit E) in clinically normal llamas. ANIMALS: 270 llamas ranging in age from < 1 month to > 15 years and grouped by age, sex, pregnancy status, and stage of gestation. Selected llamas were from 21 farms in Oregon, did not have previous health problems, and met specific health criteria on examination. PROCEDURE: Serum and blood samples were obtained and analyzed for concentrations of Ca, P, Fe, Cu, Se, Zn, and Vit E, and total iron binding capacity (TIBC) and percentage of transferrin saturation (% Sat). Mean differences by age, sex, pregnancy status, and stage of gestation, as well as all interactions, were compared to establish reference values. RESULTS: Mean values and reference ranges for most of the minerals and vitamins were similar to previously reported values. Male versus female differences were not identified for any measurements. Age was a significant variable for Ca, P, Fe, and Se concentrations, as well as Ca-to-P ratio and TIBC. Identified age-based effects were modeled by use of linear regression. Copper and Zn concentrations and % Sat did not differ as a function of age. Serum Vit E concentration was influenced by an age by sex interaction and stage of gestation. CONCLUSIONS: Age was found to be an important variable influencing many blood nutrient concentrations in healthy llamas. CLINICAL RELEVANCE: Clinical diagnosis of metabolic disease may be improved with use of age-based reference values, especially for neonates.

Aging↗
PubMed1998

Patient attitudes about mandatory reporting of domestic violence. Implications for health care professionals.

As of January 1994, California physicians are required to report to police all patients who are suspected to be victims of domestic violence. This article describes the results from a focus group study of abused women (n = 51) that explored their experiences with and perspectives on medical care. The eight focus groups included two Latina (total n = 14), two Asian (total n = 14), two African-American (total n = 9), and two Caucasian (total n = 14) groups of women who had been the victims of domestic abuse within the previous 2 years. The women were recruited through community-based organizations in the San Francisco Bay Area. With regard to physician reporting of domestic violence to police, five themes were identified: fear of retaliation by the abuser, fear of family separation, mistrust of the legal system, desire for police protection, and preference for confidentiality and autonomy in the patient-health professional relationship. Our results indicate that mandatory reporting may pose a threat to the safety and well-being of abused women and may create barriers to their seeking help and communicating with health care professionals about domestic violence.

Adult↗
PubMed1997

Characterization of guanylate kinase-associated protein, a postsynaptic density protein at excitatory synapses that interacts directly with postsynaptic density-95/synapse-associated protein 90.

The structure of central synapses is poorly understood at the molecular level. A recent advance came with the identification of the postsynaptic density-95 (PSD-95)/synapse-associated protein 90 family of proteins as important mediators of the synaptic clustering of certain classes of ion channels. By yeast two-hybrid screening, a novel protein termed guanylate kinase-associated protein (GKAP) has been isolated that binds to the GK-like domain of PSD-95 (). Here we present a detailed characterization of GKAP expression in the rat brain and report the cloning of a novel GKAP splice variant. By Northern blot, GKAP mRNAs (4, 6.5, and 8 kB) are expressed predominantly in the rat brain. By in situ hybridization, GKAP is expressed widely in neurons of cortex and hippocampus and in the Purkinje and granule cells of the cerebellum. On brain immunoblots, two prominent bands of 95 and 130 kDa are detected that correspond to products of short and long N-terminal splice variants of GKAP. Two independent GKAP antibodies label somatodendritic puncta in neocortical and hippocampal neurons in a pattern consistent with synaptic elements. Immunogold electron microscopy reveals GKAP to be predominantly postsynaptic and present at asymmetric synapses and in dendritic spines. The distribution of GKAP immunogold particles is uniform in the lateral plane of the PSD but peaks in the perpendicular axis approximately 20 nm from the postsynaptic membrane. In cultured hippocampal neurons GKAP immunoreactive puncta colocalize with the AMPA receptor subunit Glu receptor 1 but not with the GABAA receptor subunits beta2 and beta3. Thus GKAP is a widely expressed neuronal protein localized specifically in the PSD of glutamatergic synapses, consistent with its direct interaction with PSD-95 family proteins.

Amino Acid Sequence↗
PubMed1997

GKAP, a novel synaptic protein that interacts with the guanylate kinase-like domain of the PSD-95/SAP90 family of channel clustering molecules.

The molecular mechanisms underlying the organization of ion channels and signaling molecules at the synaptic junction are largely unknown. Recently, members of the PSD-95/SAP90 family of synaptic MAGUK (membrane-associated guanylate kinase) proteins have been shown to interact, via their NH2-terminal PDZ domains, with certain ion channels (NMDA receptors and K+ channels), thereby promoting the clustering of these proteins. Although the function of the NH2-terminal PDZ domains is relatively well characterized, the function of the Src homology 3 (SH3) domain and the guanylate kinase-like (GK) domain in the COOH-terminal half of PSD-95 has remained obscure. We now report the isolation of a novel synaptic protein, termed GKAP for guanylate kinase-associated protein, that binds directly to the GK domain of the four known members of the mammalian PSD-95 family. GKAP shows a unique domain structure and appears to be a major constituent of the postsynaptic density. GKAP colocalizes and coimmunoprecipitates with PSD-95 in vivo, and coclusters with PSD-95 and K+ channels/NMDA receptors in heterologous cells. Given their apparent lack of guanylate kinase enzymatic activity, the fact that the GK domain can act as a site for protein-protein interaction has implications for the function of diverse GK-containing proteins (such as p55, ZO-1, and LIN-2/CASK).

Amino Acid Sequence↗
PubMed1997

Competitive binding of alpha-actinin and calmodulin to the NMDA receptor.

The mechanisms by which neurotransmitter receptors are immobilized at postsynaptic sites in neurons are largely unknown. The activity of NMDA (N-methyl-D-aspartate) receptors is mechanosensitive and dependent on the integrity of actin, suggesting a functionally important interaction between NMDA receptors and the postsynaptic cytoskeleton. alpha-Actinin-2, a member of the spectrin/dystrophin family of actin-binding proteins, is identified here as a brain postsynaptic density protein that colocalizes in dendritic spines with NMDA receptors and the putative NMDA receptor-clustering molecule PSD-95. alpha-Actinin-2 binds by its central rod domain to the cytoplasmic tail of both NR1 and NR2B subunits of the NMDA receptor, and can be immunoprecipitated with NMDA receptors and PSD-95 from rat brain. Intriguingly, NR1-alpha-actinin binding is directly antagonized by Ca2+/calmodulin. Thus alpha-actinin may play a role in both the localization of NMDA receptors and their modulation by Ca2+.

Actins↗
PubMed1997

Activity regulates the synaptic localization of the NMDA receptor in hippocampal neurons.

We describe here a novel effect of activity on the subcellular distribution of NMDA receptors in hippocampal neurons in culture. In spontaneously active neurons, NMDA receptors were clustered at a few synaptic and nonsynaptic sites. Chronic blockade of NMDA receptor activity induced a 380% increase in the number of NMDA receptor clusters and a shift to a more synaptic distribution. This effect was reversible. The distributions of the presynaptic marker synaptophysin, the AMPA-type glutamate receptor subunit GluR1, and the putative NMDA receptor clustering protein PSD-95 were not affected by blockade. Regulation of the synaptic localization of NMDA receptors by activity may define a novel mechanism by which input controls a neuron's ability to modify its synapses.

2-Amino-5-phosphonovalerate↗
PubMed1996

Atypical pneumonia associated with ryegrass staggers in calves.

A group of 6- to 8-month-old calves developed head tremors, stiff gait, and staggering after consuming ryegrass straw that contained 3,711 micrograms of lolitrem-B/ kg. Signs were consistent with ryegrass staggers syndrome. At necropsy, all calves examined had atypical interstitial pneumonia, with marked emphysema and bullae. Infectious organisms and pneumotoxins were not identified. Experimentally, feeding the same ryegrass straw to age-matched calves induced similar neurologic signs, but did not result in pneumonic lesions. The high concentration of lolitrem-B in the straw or other, undefined factors, such as feed changes, may have contributed to the atypical interstitial pneumonia in the naturally exposed calves.

Acremonium↗
PubMed1996

Clustering of gephyrin at GABAergic but not glutamatergic synapses in cultured rat hippocampal neurons.

The molecular mechanisms underlying the establishment of a postsynaptic receptor mosaic on CNS neurons are poorly understood. One protein thought to be involved is gephyrin, a peripheral membrane protein that binds to the inhibitory glycine receptor and functions in clustering this receptor at synapses in cultured rat spinal cord neurons. We investigated the possible association of gephyrin with synapses in cultured rat hippocampal neurons, where glutamate and GABA but not glycine are the principal transmitters. Gephyrin immunoreactivity was detected in axons as well as dendrites, changing from a predominantly axonal to a more dendritic distribution with time in culture. Gephyrin staining was not distributed uniformly, but always took the form of clusters. Small clusters of gephyrin (0.2 microns 2), present throughout development, were distributed widely and not restricted to synaptic sites. Larger clusters of gephyrin (0.4-10.0 microns 2, sometimes composed of groups of small clusters), which developed in older cells, were localized to a subset of contacts between axons and dendrites. These large clusters were not present at glutamatergic synapses (marked by immunostaining for GluR1), but were closely associated with GABAergic synapses (marked by immunostaining for GABA and glutamic acid decarboxylase). These results, together with previous findings, suggest that gephyrin may function to anchor GABA and glycine receptors, but not glutamate receptors, at postsynaptic sites on central neurons. They also raise the possibility that gephyrin has additional functions, independent of its role at synapses.

Animals↗
PubMed1996

Fluconazole versus oral polyenes in the prophylaxis of immunocompromised patients: a cost-minimization analysis.

This study compares 100 mg daily fluconazole with oral polyenes four times daily in the prophylaxis of fungal infections in immunocompromised patients, to determine a cost-minimization strategy. Data was gathered through a literature survey and clinical interviews conducted in nine different UK hospitals. This was used to construct a decision tree, modelling the drug choices available to a clinician at various stages of a patient's treatment, and assigning probabilities to the different corresponding outcomes. UK cost data were fed into this model to determine the expected cost per patient of the different prophylaxis strategies. Two different patient groups were considered: chemotherapy-only patients, and bone-marrow-transplant (BMT) patients who have higher risks of fungal infection. Probabilities derived from the literature suggest that a cost-minimization strategy to manage both chemotherapy patients and BMT patients is to administer oral fluconazole, both as prophylaxis and as first line treatment, against superficial fungal infection. Probabilities gathered from clinical interviews yield similar results, suggesting that the cost-minimization strategy with chemotherapy-only patients is to administer oral polyenes as prophylaxis, and oral fluconazole in case of superficial fungal infection, while for BMT patients it is a combination of fluconazole and oral polyenes as prophylaxis, with oral fluconazole for the treatment of superficial fungal infections. Using the probabilities from the literature, the lowest cost strategies produce an expected cost of pounds 567.20 for chemotherapy-only patients, and an expected cost of pounds 804.87 for BMT patients for a course of treatment lasting from seven to 28 days. The clinical interview probabilities produce expected costs of pounds 826.48 and pounds 1529.43, respectively. Sensitivity analysis was then conducted, and it was found that in the majority of cases, using the literature probabilities, the cost-minimizing strategy remained prophylaxis with oral fluconazole. The sensitivity analysis for chemotherapy-only patients using the interview probabilities tended to favour oral polyenes as the cost-minimization strategy, whereas for BMT patients the sensitivity analysis favoured a combination of fluconazole and oral polyenes in the majority of cases. The key economic advantage of prophylaxis with fluconazole or a combination of fluconazole with oral polyenes in the prophylaxis of fungal infection in immunocompromised patients, results from the reduction of the expected cost of subsequent fungal infection among those who are most at risk.

Antifungal Agents↗
PubMed1996

Decision analysis of Helicobacter pylori eradication therapy using omeprazole with either clarithromycin or amoxicillin.

In patients with duodenal ulcer, omeprazole plus clarithromycin (OC) has achieved Helicobacter pylori eradication rates of about 80%, compared with 50% for omeprazole plus amoxicillin (OA). The drug acquisition costs for OC are 102.92 pounds sterling (pounds) compared with 38.96 pounds for OA using generic amoxicillin and 51.63 pounds using the proprietary brand 'Amoxil' (costs for 2-week regimens in 1995). The aim of this analysis was to estimate the total healthcare costs to the general practitioner (GP) of eradication therapy using a simple generalised model. Data about current practice in the UK were obtained from 502 respondents in a survey of hospital specialists and GPs. It was assumed that patients would derive no benefit from eradication therapy unless they had a duodenal ulcer, and that all OA patients received generic amoxicillin. The survey confirmed that OA was the commonest eradication therapy prescribed by UK GPs at that time. Three distinct patient groups were identified: patients with proven duodenal ulcer who were already receiving maintenance treatment with a histamine H2 receptor antagonist, and new patients with dyspepsia who were subdivided into those aged above or below 45 years. Patients receiving maintenance treatment for a duodenal ulcer would be prescribed eradication therapy by their GP without further endoscopy. If dyspepsia recurred after eradication therapy, they would be referred to a gastroenterologist, who would perform an endoscopy to confirm the recurrence of ulceration. In this model, the expected total healthcare costs (i.e. the costs of drug acquisition and subsequent treatment when required) following prescription of eradication therapy were lower for OC (157 pounds) than for OA (173 pounds). New patients aged over 45 years would be referred for endoscopy because of the risk that dyspepsia might be the initial presentation of gastric cancer. If duodenal ulceration was found, eradication therapy would be prescribed and, if dyspepsia remained or recurred, the patient would be referred back to the gastroenterologist. In this case, it was considered unlikely that a further endoscopy would be performed. Thus, the healthcare costs associated with failure of eradication in these patients were less than for patients on maintenance treatment, and the expected total healthcare costs were higher for OC (349 pounds) than for OA (335 pounds). Finally, a new patient aged under 45 years with dyspepsia would have eradication therapy prescribed on the basis of a clinical diagnosis of duodenal ulcer plus serological evidence of infection with H. pylori. Continuation or recurrence of dyspepsia would result in referral to a gastroenterologist, who would perform an endoscopy. The total expected healthcare costs were higher for OC (253 pounds) than for OA (251 pounds). The cost effectiveness of OA was sensitive to changes in the default costs (i.e. the average costs from the survey used in the decision analysis), particularly in patients < 45 years old. In these patients, OC would become the cheaper option if amoxicillin were prescribed by brand name instead of in generic form. In this patient group, the outcome was crucially dependent on the accuracy of the clinical diagnosis of duodenal ulcer; if this was at least 60%, then OC would be the cheaper regimen. Overall, the model clearly shows that the higher drug cost of OC is likely to be substantially offset by savings in other healthcare costs. If the direct healthcare costs of OC are higher than OA, then the decision maker must consider the indirect and intangible costs associated with failure of eradication therapy.

Adult↗
PubMed1996

Vitamin E supplementation and stress affect tissue alpha-tocopherol content of beef heifers.

The effect of stress on tissue alpha-tocopherol was investigated in 16 crossbred heifers fed a corn/corn silage-based diet. For 28 d, eight heifers (379 +/- 10 kg BW) received a dietary supplement of 1,000 IU of dl-alpha-tocopheryl acetate, whereas the controls (375 +/- 10 kg BW) received no supplemental vitamin E. Tissue samples of plasma, red blood cells, liver, trapezius, and longissimus muscles and subcutaneous fat immediately dorsal to each muscle were taken on d 1 for determination of alpha-tocopherol concentration. On d 2 through 4 each heifer was restricted to 2.61 kg of grass hay and allowed water. On d 5, 6, and 7 no feed or water was given, 100 IU of ACTH and .0024 mg of epinephrine/kg BW were given every 8 h, and biopsies for alpha-tocopherol content were again taken on d 7. The stress reduced (P < .01) mean BW, increased (P < .01) serum cortisol, creatine kinase, and urea. After stress, supplemental vitamin E reduced (P < .13) the increase in creatine kinase relative to that in heifers not supplemented with vitamin E. Stress also increased (P < .04) serum Se in heifers fortified with the vitamin E. Alpha-tocopherol content of plasma, red blood cells, liver, and subcutaneous fat dorsal to the trapezius muscle was increased (P < .01) by supplemental vitamin E. The stress treatment reduced (P < .01) alpha-tocopherol content of plasma in those fed the vitamin E and increased it (P < .05) in the nonsupplemented vitamin E-deficient heifers. Stress also decreased red blood cell (P < .01) and liver (P < .05) alpha-tocopherol content in cattle supplemented with vitamin E. Tissue alpha-tocopherol concentrations were reduced by stress only when a diet adequate in vitamin E was fed. In addition, in most sampled tissues, stress did not affect alpha-tocopherol concentrations.

Adipose Tissue↗
PubMed1995

Preferential addition of newly synthesized membrane protein at axonal growth cones.

The addition of plasma membrane proteins to a growing axon could occur by preferential insertion at the tip (the growth cone), by uniform insertion along the axon, or by insertion at the cell body and bulk flow along the axon. To differentiate between these possibilities we used a defective herpesvirus vector to express an exogenous protein, the lymphocyte transmembrane protein CD8 alpha, in cultured rat hippocampal neurons. The newly synthesized protein first appeared on the axonal surface almost exclusively at the growth cone. Preferential addition at the growth cone was also observed in minor processes (immature dendrites), but not in mature dendrites. Over several hours, CD8 alpha reached a uniform distribution over the entire neuronal surface, presumably by diffusion within the membrane and possibly endocytic recycling. As well as providing materials for axonal growth, the selective addition of membrane vesicles at the growth cone may contribute to the polarized distribution of axonal surface molecules.

Animals↗
PubMed1995

Market structure and conduct in the pharmaceutical industry.

The pharmaceutical industry offers the world's population alleviation and cure from a variety of medical conditions, while also contributing to the economic performance of many countries. The potential importance of these characteristics have made the industry an area of intense debate, criticism and praise. This review aims to clarify some of the arguments surrounding the industry on both sides of the fence, and by doing so, to provide a balanced introduction on which opinions may be made.

Economics, Pharmaceutical↗
PubMed1994

Selective clustering of glutamate and gamma-aminobutyric acid receptors opposite terminals releasing the corresponding neurotransmitters.

Several immunocytochemical and physiological studies have demonstrated a concentration of neurotransmitter receptors at postsynaptic sites on neurons, but an overall picture of receptor distribution has not emerged. In particular, it has not been clear whether receptor clusters are selectively localized opposite terminals that release the corresponding neurotransmitter. By using antibodies against the excitatory glutamate receptor subunit GluR1 and the inhibitory type A gamma-aminobutyric acid (GABA) receptor beta 2/3 subunits, we show that these different receptor types cluster at distinct postsynaptic sites on cultured rat hippocampal neurons. The GABAA receptor beta 2/3 subunits clustered on cell bodies and dendritic shafts opposite GABAergic terminals, whereas GluR1 clustered mainly on dendritic spines and was associated with glutamatergic synapses. Chronic blockade of evoked transmitter release did not block receptor clustering at postsynaptic sites. These results suggest that complex mechanisms involving nerve terminal-specific signals are required to allow different postsynaptic receptor types to cluster opposite only appropriate presynaptic terminals.

Action Potentials↗
PubMed1994

Reduced malignancy of ras-transformed NIH 3T3 cells expressing antisense osteopontin RNA.

Osteopontin (OPN) is a secreted, calcium-binding phosphoprotein that frequently has been associated with the transformed phenotype. To clarify the function of OPN in tumor cells, we designed experiments to: (a) express antisense OPN RNA in murine PAP2 cells (metastatic, ras-transformed NIH 3T3 cells) and (b) examine the effects of antisense OPN expression on the tumorigenic and metastatic properties of the cells. PAP2 cells were transfected with pNMH-asOPN, an inducible, mammalian expression vector that can generate antisense OPN RNA complementary to the OPN mRNA. Two clones have been identified that expressed antisense OPN RNA in vitro. While reduced OPN protein secretion was not detected when the cells were grown in vitro, the in vivo expression of antisense OPN RNA was associated with reduced tumorigenicity. Tumors that did arise, with greatly extended lag time, had lost expression of antisense OPN RNA in vivo, suggesting that antisense OPN RNA expression was associated with reduced tumorigenicity of these cells.

3T3 Cells↗