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L R Jones

Publications and source records attributed to L R Jones.

At least 163 records · Page 9Linked to original sources
PubMed1984

High molecular weight proteins in cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles bind calmodulin, are phosphorylated, and are degraded by Ca2+-activated protease.

A unique set of high molecular weight proteins was identified in junctional sarcoplasmic reticulum (SR) vesicles isolated from both cardiac muscle and skeletal muscle. These high Mr proteins were not present in free SR vesicles isolated from either tissue, nor were they observed in purified sarcolemmal fractions. The junctional SR high Mr proteins migrated as doublets in sodium dodecyl sulfate-polyacrylamide gels and exhibited apparent Mr values between 290,000 and 350,000. The high Mr proteins bound calmodulin; they were the principal proteins labeled in the cardiac and skeletal muscle SR subfractions by azido-125I-calmodulin. The high Mr proteins were also substrates for an endogenous Ca2+-calmodulin-dependent protein kinase activity, as well as exogenously added catalytic subunit of cAMP-dependent protein kinase. In addition, the junctional SR high Mr proteins were the major SR proteins degraded by a Ca2+-activated protease purified from smooth muscle. Control experiments verified the separation of junctional SR vesicles and free SR vesicles from both muscle types. Junctional SR vesicles were enriched in calsequestrin, and they exhibited Ca2+ uptake which was stimulated up to 10-fold by either ryanodine or ruthenium red. Free SR vesicles were deficient in calsequestrin and were insensitive to these two agents. Localization of the cardiac and skeletal muscle high Mr proteins to the junctional SR, coupled with demonstration of their nearly identical biochemical properties, suggests that the proteins are homologous and are likely to have similar functions in both types of striated muscle.

Animals↗
PubMed1984

Phosphorylation-induced mobility shift in phospholamban in sodium dodecyl sulfate-polyacrylamide gels. Evidence for a protein structure consisting of multiple identical phosphorylatable subunits.

Phosphorylation of purified phospholamban isolated from canine cardiac sarcoplasmic reticulum vesicles decreased the electrophoretic mobility of the protein in sodium dodecyl sulfate (SDS)-polyacrylamide gels. Different mobility forms of phospholamban in SDS gels were visualized both by direct protein staining and by autoradiography. Unphosphorylated phospholamban migrated with an apparent Mr = 25,000 in SDS gels; maximal phosphorylation of phospholamban by cAMP- or Ca2+-calmodulin-dependent protein kinase increased the apparent Mr to 27,000. Partial phosphorylation of phospholamban by either protein kinase gave intermediate mobility forms of molecular weights between 25,000 and 27,000, suggesting that more than one phosphorylation site was present on the holoprotein for each activity. Boiling of phospholamban in SDS dissociated the holoprotein into an apparently homogeneous class of low molecular weight "monomers." Only two mobility forms of monomeric phospholamban were observed in SDS gels after phosphorylation by cAMP-dependent protein kinase, corresponding to 9-kDa dephospho- and 11-kDa phosphoproteins. All of the 9-kDa protein could be phosphorylated and converted into the 11-kDa mobility form, suggesting the presence of only one site of phosphorylation on a single type of monomer for cAMP-dependent protein kinase. Simultaneous phosphorylation of monomeric phospholamban by cAMP-dependent protein kinase and Ca2+-calmodulin-dependent protein kinase gave an additional mobility form of the protein, suggesting that different sites of phosphorylation were present for each activity on each monomer. Incomplete dissociation of the holoprotein by boiling it in a relatively low concentration of SDS facilitated the detection of five major mobility forms of the protein in SDS gels, and the mobilities of all of these forms were decreased by phosphorylation. We propose that the high molecular weight form of phospholamban is a multimer of electrophoretically indistinguishable monomers, each of which contains a different phosphorylation site for cAMP-dependent protein kinase activity and Ca2+-calmodulin-dependent protein kinase activity. Phosphorylation of phospholamban at multiple sites is responsible for the various mobility forms of the holoprotein detected in SDS-polyacrylamide gels.

Adenosine Triphosphatases↗
PubMed1984

Forensic training in family practice residencies.

Knowledge and skill in forensic medicine are important in primary care not only for defensive purposes but also because of potential therapeutic value in patient care. The major role in future mental health services envisioned for primary care physicians makes such training especially important. A national survey of family practice residency programs reveals that 47 percent of programs do not address forensic aspects of medical practice. A model forensic medicine curriculum is described that would require minimal adjustment of existing programs. The need for inclusion of forensically qualified clinicians in training programs for primary care physicians is evident.

Curriculum↗
PubMed1983

Rapid purification of calsequestrin from cardiac and skeletal muscle sarcoplasmic reticulum vesicles by Ca2+-dependent elution from phenyl-sepharose.

Treatment of cardiac or skeletal muscle sarcoplasmic reticulum vesicles with 0.1 M sodium carbonate selectively extracts both the Ca2+-binding protein calsequestrin and the two "intrinsic glycoproteins," while leaving the Ca2+-dependent ATPase membrane bound. Phenyl-Sepharose chromatography in the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and high salt (0.5 M NaCl) readily fractionates these solubilized proteins into a Ca2+-elutable fraction, which contains purified calsequestrin, and a low ionic strength elutable fraction, which contains one of the two intrinsic glycoproteins. Elution of calsequestrin from phenyl-Sepharose occurs near 1 mM Ca2+. Copurifying with calsequestrin are an homologous set of high molecular weight proteins, which like calsequestrin stain blue with Stains-All. These proteins are present in trace amounts and do not correspond to any sarcoplasmic reticulum proteins previously identified. Elution of calsequestrin from phenyl-Sepharose is consistent with the Ca2+-binding protein losing its hydrophobic character in the presence of millimolar Ca2+. This behavior is converse to that observed for several calmodulin-like proteins, which are eluted from hydrophobic gels in the presence of EGTA. The high yield and purity of calsequestrin prepared by this method makes possible a unique system for studying what may be a distinct class of Ca2+-binding proteins.

Animals↗
PubMed1983

The cardiac beta-adrenergic receptor. Structural similarities of beta 1 and beta 2 receptor subtypes demonstrated by photoaffinity labeling.

The beta-adrenergic receptor photoaffinity ligand p-azido-m-[125I]iodobenzylcarazolol has been used to covalently label the beta 1 and beta 2 adrenergic receptor binding subunits present in left ventricular myocardial membranes derived from mammalian (including human) and nonmammalian species. Covalent incorporation of the photoaffinity ligand into membrane proteins was followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the case of the human, canine, porcine, rabbit, and rat left ventricle, all of which contain predominantly or exclusively beta 1-adrenergic receptors, two peptides of Mr approximately equal to 62,000 (major component) and Mr approximately equal to 55,000 (minor component) were specifically labeled and visualized by autoradiography. Photoincorporation into these two bands could be blocked with the appropriate drugs to display a beta 1-adrenergic receptor pharmacological specificity. Simultaneous sodium dodecyl sulfate-polyacrylamide gel electrophoresis of samples from each species revealed that all of the Mr = 62,000 peptides co-migrated suggesting similarity in the beta 1-adrenergic receptor binding subunit peptides in all of these species. The minor component Mr approximately equal to 55,000 appears to be a proteolytic degradation product of the Mr = to 62,000 peptide. Its formation could be decreased by proteinase inhibitors. This suggests that the heterogeneity of the labeling pattern observed in mammalian tissues in this and previous studies may be the result of proteolytic degradation of the receptor subunit which occurs during membrane preparation. Photoaffinity labeling of frog ventricular membranes which contain predominantly beta 2-adrenergic receptors also revealed two peptides of Mr approximately equal to 62,000 (major component) and 55,000 (minor component) with the pharmacological selectivity of a beta 2-adrenergic receptor. These data suggest marked similarities in the beta 1- and beta 2-adrenergic receptor binding subunits of different species and suggest that the pharmacological subtype might be determined by the detailed structure, i.e. amino acid sequence, at the ligand binding sites of the receptor peptide.

Adult↗
PubMed1983

Enrichment, solubilization, and partial characterization of digitonin-solubilized muscarinic receptors derived from canine ventricular myocardium.

The subcellular distribution of cardiac muscarinic receptors was defined in canine ventricular myocardium, and receptors were solubilized from subcellular fractions enriched in muscarinic receptor content. The subcellular location of muscarinic receptors in cardiac tissue was determined by measurement of the distribution of [3H](+/-)quinuclidinyl benzilate-binding activity in particulate fractions isolated from canine ventricular myocardium. Based upon excellent correlation between [3H](+/-)quinuclidinyl benzilate binding and activity of the sarcolemmal Na+,K+-ATPase throughout the subcellular fractions, muscarinic receptors appeared to be localized to sarcolemma in canine ventricular myocardium. Therefore, membrane fractions enriched in sarcolemma were used as a source of cardiac muscarinic receptors for solubilization. Treatment of membrane vesicle fractions with digitonin (0.6%) resulted in solubilization of [3H](+/-)quinuclidinyl benzilate-binding activity with an extraction yield of 25-35%. Criteria of pharmacological specificity and stereospecificity established the identity of the solubilized binding activity of muscarinic receptors. Solubilization of muscarinic receptors was documented by demonstration of hydrodynamic behavior consistent with molecularly dispersed material. Upon glycerol gradient centrifugation, digitonin-solubilized muscarinic receptors from cardiac tissue sedimented with an apparent sedimentation coefficient of 9S. Pharmacological characterization of the digitonin-solubilized receptors revealed 8- to 39-fold reductions in affinities for muscarinic antagonists compared to the affinities exhibited by receptors in the membrane-bound state. Substantially greater reductions in agonist affinities (reduction of at least 700-fold for all agonists studied) suggested selective loss of ability of the digitonin-solubilized receptors to exhibit high affinity agonist interactions. In contrast to membrane-bound receptors, digitonin-solubilized receptors also demonstrated a loss of guanine nucleotide regulation, as well as steep agonist:radioligand competition curves with slope factors of 1.0, suggesting a homogeneous population of agonist-binding sites. Interpreted within the context of a model of state interconversion for membrane-bound receptors, the results suggested that either muscarinic receptors of a single state were selectively solubilized, or that solubilization induced conversion of all receptors to a single low affinity state, possibly by removal of constituents necessary for assumption of a high affinity agonist conformation.

Animals↗
PubMed1982

Characterization of the intrinsic cAMP-dependent protein kinase activity and endogenous substrates in highly purified cardiac sarcolemmal vesicles.

The intrinsic cAMP-dependent protein kinase activity of highly purified cardiac sarcolemmal vesicles was characterized. The sarcolemmal protein kinase was specifically activated by cAMP. Binding of cAMP to the kinase was saturable and occurred exclusively to a protein of Mr = 55,000 intrinsic to the vesicles. This binding of cAMP to the sarcolemmal vesicles caused a selective release of catalytic activity from the membranes, which was capable of phosphorylating several endogenous sarcolemmal substrates as well as one additional substrate, which was also identified in purified vesicles of cardiac sarcoplasmic reticulum. Unmasking experiments conducted with the ionophore alamethicin demonstrated that the protein kinase activity and its endogenous sarcolemmal substrates were localized on the inner, cytoplasmic surfaces of the vesicles, and, furthermore, suggested that at least 75% of the vesicles were right side out. The major protein substrates phosphorylated in the sarcolemmal fraction exhibited apparent molecular weights of 21,000 and 8,000, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Heating the membranes in the presence of sodium dodecyl sulfate prior to electrophoresis completely converted the 21,000-dalton substrate into the form of higher mobility, suggesting that the two substrates were, in fact, identical proteins. This was supported by the observation that both substrates exhibited identical pI values of approximately 6.7. Although present in the sarcolemmal fraction, these two substrates were not localized exclusively to sarcolemmal membranes. The same two substrates were present in 3-fold higher content in purified cardiac sarcoplasmic reticulum vesicles. Moreover, although phosphorylation of all other sarcolemmal proteins in right side out vesicles by exogenously added protein kinase was increased 4-fold or greater by alamethicin, phosphorylation of the substrates of Mr = 21,000 and 8,000 was not altered appreciably by the ionophore. The results suggest that these two major substrates identified in the sarcolemmal preparations are not intrinsic sarcolemmal proteins.

Adenosine Triphosphatases↗
PubMed1982

Mental health services in primary health care: yet another era of disillusionment?

Amid the ruins of a mental health care delivery system chronically beleaguered by inadequate planning, financial support, or public consensus, efforts to integrate mental health services with the health services of primary care physicians are considered the best hope to improve access to mental health care. Despite a high prevalence of mental disorder in their patients, most primary care physicians do not recognize, formally diagnose, or treat emotional disorders. Significant personal, professional, and economic issues may compromise effective mental health care delivery in underserved areas, especially poor access to continued medical education and professional support resources. The thrust to provide primary care manpower is at the expense of core health professional training in other needed specialties, including psychiatry. Comprehensive planning for mental health services to underserved populations must consider the shortage and maldistribution of psychiatrists, requirements for professional team and liaison support for effective mental health care delivery by primary care physicians, access to continuing education, and reimbursement for appropriate mental health services. Increasing the mental health role of primary care physicians may otherwise amount to no more than a shift in the locus of expected care delivery.

Family Practice↗
PubMed1982

Medical illness in psychiatric patients: barriers to diagnosis and treatment.

We report a study of 102 consecutive admissions to the acute medical care unit of a large psychiatric hospital. The study was designed to investigate the epidemiology and the barriers to diagnosis and treatment of medical illness among female psychiatric inpatients. The majority of the patients were transferred to the unit because of nonspecific changes in physical condition or for behavioral differences. More than 70% of the patients were unable to communicate adequately with the physician. Ninety-two percent of the sample were found to have at least one with an average of three previously undiagnosed physical diseases not predicted by their symptoms on referral. We advocate a high index of suspicion of physical disease in the psychiatric population, and recommend an aggressive multidisciplinary diagnostic and therapeutic approach.

Adolescent↗
PubMed1982

Mental health training in family practice residency programs.

This paper presents a national survey of psychiatric and behavioral science training in approved family medicine residency programs. A 64 percent response rate resulted in data describing residency programs approved by the American Academy of Family Physicians: the residents, faculty, and curriculum content, and the teaching-learning format utilized in mental health training. Though improvement in the mental health services of primary care providers is a major health issue, training is typically provided by part-time or volunteer faculty; less than 5 percent of the total faculty are full-time psychiatrists. Critical elements of effective curriculum design and content that are inadequate or omitted are discussed.

Curriculum↗
PubMed1981

Biochemical evidence for functional heterogeneity of cardiac sarcoplasmic reticulum vesicles.

Two subpopulations of cardiac sarcoplasmic reticulum vesicles were resolved functionally, based on their sensitivities to the drug ryanodine. These two subpopulations of sarcoplasmic reticulum vesicles, termed ryanodine-sensitive and ryanodine-insensitive, were separated by preloading crude cardiac microsomes with Ca2+ oxalate in the presence of ATP, followed by sucrose density gradient centrifugation. Ryanodine-insensitive vesicles accumulated most of the Ca2+ oxalate during the preload, and constituted the densest subfraction recovered from the sucrose gradient. These ryanodine-insensitive vesicles exhibited the highest density of Ca2+ pumps, and accounted for 10 to 15% of the total protein in crude cardiac microsomes. Ryanodine-insensitive vesicles continued to transport substantial amounts of Ca2+ after isolation. Ryanodine-sensitive vesicles accumulated negligible Ca2+ during the preload, and were recovered from the lower density regions of the sucrose gradient. On a milligrams of protein basis, these vesicles were present in 7-fold excess over ryanodine-insensitive vesicles. Ryanodine-sensitive vesicles transported low amounts of Ca2+ under normal incubation conditions, but 3 X 10(-4) M ryanodine strikingly increased their Ca2+ uptake 5- to 10-fold. Ca2+ uptake by ryanodine-sensitive vesicles was uniquely regulated by Ca2+ ion concentration. Elevation of the ionized Ca2+ concentration from 2 to 4 microM increased Ca2+ uptake by these vesicles greater than 5-fold, but had no effect on their Ca2+-dependent ATPase activity. These ryanodine- and Ca2+ concentration-dependent effects were apparent for only ryanodine-sensitive vesicles. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed distinct differences in polypeptide staining between ryanodine-sensitive and ryanodine-insensitive vesicles, confirming by an independent method that the two populations of vesicles were different. These data provide the first biochemical evidence for functional and structural heterogeneity of cardiac sarcoplasmic reticulum vesicles.

Adenosine Triphosphate↗
PubMed1981

Membrane localization of myocardial type II cyclic AMP-dependent protein kinase activity.

Crude cardiac membrane vesicles were separated into subfractions of sarcolemma and sarcoplasmic reticulum. The subfractions were used to determine the origin and type of cyclic AMP-dependent protein kinase activity present in myocardial membranes. A cyclic AMP-binding protein of molecular weight 55,000 was covalently labeled with the photoaffinity probe 8-azido adenosine 3',5'-mono[32P]phosphate, and found to copurify with the (Na+ + K+)-ATPase activity of sarcolemma, and away from the (Ca2+ + K+)-ATPase activity of sarcoplasmic reticulum. Endogenous cyclic AMP-dependent protein kinase activity also copurified with sarcolemma. Protein substrates phosphorylated by cyclic AMP-dependent protein kinase activity had apparent molecular weights of 21,000 and 8000 and were present in both sarcolemma and sarcoplasmic reticulum. However, while addition of cyclic AMP alone resulted in phosphorylation of sarcolemma proteins, both cyclic AMP and exogenous, soluble cyclic AMP-dependent kinase were required for phosphorylation of sarcoplasmic reticulum proteins. Addition of the calcium-binding protein, calmodulin, to either sarcolemma or sarcoplasmic reticulum resulted in phosphorylation of the 21,000 and 8000-dalton proteins, as well. The results suggest that cardiac sarcolemma contains an intrinsic type II cyclic AMP-dependent protein kinase activity that is not present in sarcoplasmic reticulum. On the other hand, Ca2+- and calmodulin-dependent protein kinase activity is present in both sarcolemma and sarcoplasmic reticulum.

Adenosine Triphosphatases↗
PubMed1981

A curriculum for the psychiatric training of family physicians.

This curriculum is designed with the expectation of considerable involvement by family physicians in mental health care delivery. Psychiatry education is initiated early, before attitudes become fixed, to emphasize self-reliance and mental health problem-solving skills, to make the learning experience longitudinal, and to integrate psychiatric training into the rest of the residents' curriculum. Seminars, clinical experience, and liaison with a mental health team are all utilized in the training design. The effectiveness of this training program will be studied over time to obtain needed data about the best teaching model for the mental health role of the family physician.

Curriculum↗