Inappropriate prescribing of intravenous fluid in adult inpatients-a literature review of current practice and research.

WHAT IS KNOWN AND OBJECTIVE: It is known that mismanagement of intravenous (IV) fluid therapy may cause serious complications. The 2013 NICE guideline on intravenous fluid therapy in hospitalized adults also emphasizes the importance of appropriate prescribing of IV fluid. So far, no systematic review of the incidence and types of inappropriate prescribing of IV fluid has been conducted. Therefore, this study was undertaken to review the research literature on inappropriate prescribing of IV fluid in adult patients and develop corresponding strategies for improving practice. METHODS: A comprehensive literature search was performed. Critical appraisals were conducted on the articles drawn from the search, and an analysis was performed on the results. RESULTS AND DISCUSSION: Incorrect volumes and types of IV fluids prescribed, classified as misprescribing, was the most common type of inappropriate prescribing. Commonly, patients on IV fluid therapy were prescribed a greater volume of fluid and amount of sodium in excess of normal requirements. Doctors did not always check the body weight, serum electrolyte level and serum creatinine before prescribing IV fluid for patients. The other common type of inappropriate prescribing was incomplete/incorrect prescription writing. These common inappropriate prescribing of IV fluid could be caused by insufficient knowledge and training of the prescribers. In addition, the ignorance of the importance of IV fluid prescribing also contributed to this behaviour. WHAT IS NEW AND CONCLUSION: There is an urgent need to make doctors aware of these problems and enhance appropriate training on IV fluid prescribing, especially on the appropriate volume and amount of electrolytes. Pharmacists could exert a role in reviewing the fluid prescription chart for improving clinical practice.

The mechanism of protein kinase C activation.

Protein kinase C (PKC) consists of a family of closely related enzymes highly concentrated in the CNS. These enzymes respond to the second messengers calcium (Ca2+) and diacylglycerol (DAG), to express their activities at membrane locations. Each member of this enzyme family displays distinct biochemical characteristics and is enriched in different cellular and subcellular locations. Activation of PKC in the nervous system has been implicated in the regulation of neurotransmitter release, ion channels, growth and differentiation, and neural plasticity. It is believed that an increase in the intracellular concentration of Ca2+ triggers the association of a group of PKC isozymes with the membrane where DAG interacts with PKC to stimulate the enzyme activity. Stimulation of PKC at the cellular membrane is, therefore, dependent upon the duration and magnitude of the DAG signal. The association of PKC with the membrane may also lead to a conversion of the enzyme into an effector-independent form for a sustained activation after the Ca2+ and DAG signals dissipate. Activation of PKC results in the phosphorylation of cellular proteins; however, the physiological substrates of this enzyme in the nervous system are still poorly characterized.

Role of protein kinase C in phosphorylation of vinculin in adriamycin-resistant HL-60 leukemia cells.

In response to phorbol esters such as 12-O-tetradecanoylphorbol-13-acetate (TPA), HL-60 cells differentiate to macrophage-like cells and exhibit the ability to phosphorylate vinculin in vitro. Adriamycin-resistant HL-60 (HL-60/ADR) cells similarly demonstrate this characteristic without prior treatment with TPA. Since protein kinase C (PK-C) is a cellular TPA receptor, we have examined the role of this enzyme in the inherent ability of HL-60/ADR cells to phosphorylate vinculin. DEAE-cellulose chromatography of cell extracts revealed that HL-60/ADR cells contained 2-fold more PK-C than did the parental cell line. All PK-C activity was found in the cytosol of wild type HL-60 cells, whereas 85% of PK-C activity was cytosolic and 15% was membrane-bound in HL-60/ADR cells. After a 2-day treatment with 10 nM TPA, PK-C activity was reduced 80-90% in both cell lines regardless of its intracellular distribution. Immunoblotting of cell extracts from HL-60/ADR cells or HL-60 cells following treatment with TPA revealed increased levels of a 52-kDa species of similar mass to M-kinase. Coincident with these changes after TPA treatment was a reduction in Ca2+ and phospholipid-independent phosphorylation of vinculin in vitro in extracts from HL-60/ADR cells, whereas HL-60 cells exhibited an elevation of this phosphoprotein. The phosphorylation of vinculin in TPA-treated HL-60 cells or untreated HL-60/ADR cells was blocked by antibodies to protein kinase C. These results suggest that it is not the absolute level of protein kinase C but rather the proteolytic activation of PK-C to a Ca2+ and phospholipid-independent form which is associated with the utilization of vinculin as an endogenous substrate.

Phosphorylation of MARCKS, neuromodulin, and neurogranin by protein kinase C exhibits differential responses to diacylglycerols.

Diacylglycerols (DG) derived from brain phosphatidylinositol (PI) and phosphatidylcholine (PC) and synthetic 1,2-dioleoylglycerol (diC18:1) and 1,2-dioctanoylglycerol (diC8) were tested for their efficacy in stimulating PKC-catalyzed phosphorylation of three physiological substrates in the brain, namely, MARCKS, neuromodulin (Nm), and neurogranin (Ng). The A0.5 of these DGs for PKC were variable dependent on the protein substrates; the values were lowest with MARCKS and highest with Ng. With Ng as a substrate the A0.5 of these DGs for PKC gamma were PI- and PC-DGs < diC18:1 < diC8. Both PI- and PC-DGs, in spite of their differences in unsaturated fatty acids content, were similarly effective in stimulating PKC. Since the phosphorylation of MARCKS, as compared to those of Nm and Ng, has the lowest A0.5 with the various DGs, it seems that among these three PKC substrates MARCKS is most readily phosphorylated by PKCs following DG formation in vivo.

Glycogen synthase (casein) kinase-1: tissue distribution and subcellular localization.

The distribution of glycogen synthase (casein) kinase-1 (CK-1) among different rat tissues and subcellular fractions was investigated. Using casein, glycogen synthase and phosphorylase kinase as substrates, CK-1 activity was detected in kidney, spleen, liver, testis, lung, brain, heart, skeletal muscle and adipose tissue. The distribution of CK-1 among different subcellular fractions of rat liver was; cytosol (72.1%), microsome (17.6%), mitochondria (9.6%) and nuclei (0.7%). CK-1 from rat tissues was shown to have a similarly wide substrate specificity as highly purified CK-1 from rabbit skeletal muscle. Such wide substrate specificity and distribution among different mammalian tissues and subcellular organelles indicate that CK-1 may be involved in the regulation of diverse cellular functions.

Catalytic fragment of protein kinase C exhibits altered substrate specificity toward smooth muscle myosin light chain.

Smooth muscle myosin light chain (LC) can be phosphorylated by myosin light chain kinase (MLCK) at Ser19 and Thr18 and by protein kinase C (PKC) at Thr9 and Ser1 or Ser2 under the in vitro assay conditions. Conversion of PKC to the spontaneously active protein kinase M (PKM) by proteolysis resulted in a change in the substrate specificity of the kinase. PKM phosphorylated both sets of sites in LC recognized by MLCK and PKC as analyzed by peptide mapping analysis. The PKM-catalyzed phosphorylation of these sites was not greatly affected by a MLCK inhibitor, ML-9, nor by the activators of MLCK, Ca2+ and calmodulin.

Phosphorylation of smooth muscle myosin light chain by five different kinases.

Phosphorylation of the 20 kDa myosin light chain from smooth muscle by five different kinases was investigated. Three of the kinases (myosin light chain kinase, phosphorylase kinase, and cAMP-dependent protein kinase) phosphorylate serine residues, the fourth (casein-kinase-2) mainly threonine, and the fifth (glycogen synthase (casein) kinase-1) both serine and threonine. Isoelectric focusing analyses of 32P-labelled chymotryptic peptides indicate that phosphorylase kinase and cAMP-dependent protein kinase phosphorylate the same site as myosin light chain kinase. However, both casein kinase-2 and glycogen synthase (casein) kinase-1 phosphorylate different sites.

Identification of a nuclear protein binding element within the rat brain protein kinase C gamma promoter that is related to the developmental control of this gene.

Protein kinase C gamma (PKC gamma) is a brain-specific isozyme expressed at a high level in the adult but not in the fetal or newborn rat. At least seventeen nuclear protein binding sites within the 5'-flanking region extending from -1612 to +243 had been identified by DNase I footprinting analysis and gel mobility shift assays. Among them, one site, GAATTAATAGG, at -669 to -679 is protected from DNase I digestion by nuclear protein from newborn but not from the adult rat brain. The levels of this binding protein, as determined by gel mobility shift assay, were found inversely related to the levels of PKC gamma in rat brain at different stages of development. These results suggest that this particular binding site may participate in the developmental regulation of PKC gamma gene.

Phosphorylation of magainin-2 by protein kinase C and inhibition of protein kinase C isozymes by a synthetic analogue of magainin-2-amide.

Magainins are a family of antimicrobial peptides present in the skin extracts of Xenopus laevis. Both magainin-1 and -2 do not have any significant effect on the activity of protein kinase C (PKC). Magainin-2 was found to be readily phosphorylated by PKC to 0.5 mol 32P/mol of peptide. Neither magainin-1, which has a sequence of S8AGK and not S8AKK as in the case of magainin-2, nor the magainin-2 analogue with substitution of Ala for Ser8 was phosphorylated by the kinase, suggesting that Ser8 is the phosphorylation site of magainin-2. One synthetic analogue of magainin, designated magainin B, which has a greater tendency for alpha-helix formation in non-aqueous environment than the parent peptide resulting from substitution of Ser8, Gly13, and Gly18 with Ala in magainin-2-amide, is a potent inhibitor of PKC. This peptide inhibits all three PKC isozymes with IC50 less than 20 microM. Magainin B also inhibits the binding of [3H]phorbol 12,13-dibutyrate to the kinase. These results suggest that magainin-2 may be modified by PKC through phosphorylation and that certain synthetic analogues of magainins may be used as inhibitors of PKC.

Reduced protein kinase C immunoreactivity and altered protein phosphorylation in Alzheimer's disease fibroblasts.

Abnormal protein kinase C levels and protein kinase C-dependent phosphorylation are biochemical alterations in brain tissue obtained from patients with Alzheimer's disease. Because many biochemical and biophysical abnormalities are found in peripheral tissues of patients with Alzheimer's disease, we studied protein kinase C levels and the in vitro phosphorylation of proteins under protein kinase C-activating conditions in fibroblasts derived from patients with Alzheimer's disease. The concentration of protein kinase C-like immunoreactivity was reduced in Alzheimer's disease samples, although the protein kinase C activity determined by the phosphorylation of exogenous histone was not. The degree of in vitro phosphorylation of an Mr 79,000 protein in the presence of protein kinase C activators was less in Alzheimer's disease than in control fibroblast cytosol, and a reduction was more prominent in cases of familial Alzheimer's disease than in sporadic Alzheimer's disease. Therefore, the aberrant phosphorylation mediated by protein kinase C is found not only in the brain but also in fibroblasts.

How is protein kinase C activated in CNS.

Protein kinase C (PKC) enzyme family consists of the Ca(2+)-dependent and -independent subgroups of phospholipid/diacylglycerol (DAG)-stimulated serine/threonine protein kinases. These enzymes exhibit distinct cellular and subcellular localizations in CNS and subtle differences in their biochemical characteristics and substrate specificities. It is believed that each of these isoenzymes respond differently to different input signals. However, detailed mechanism for the functioning of these enzymes in vivo is largely unknown; this is in part due to the absence of specific activator, inhibitor, or substrate for each of these enzymes. Recent advances in biochemical, biophysical, and molecular characterizations have defined certain structural features important to confer the stimulatory responses of these enzymes to Ca2+, DAG or phorbol ester, and Zn2+; other features important for the binding of anionic phospholipids, Ca2+/phospholipid complexes, and cis-unsaturated fatty acids have not yet been characterized. Activation of PKC requires the increase in [Ca2+]i and DAG and/or cis-unsaturated fatty acids. Ca2+ promotes the interactions of the Ca(2+)-dependent subgroup of PKCs with membrane phosphatidylserine (PS) and the enzymes become partially active when simultaneously associated with phosphatidylinositol 4,5-bisphosphate or fully active when DAG is available. Free fatty acids such as arachidonic acid, generated by the activation of phospholipase A2, could synergize with DAG to activate the enzyme maximally. The Ca(2+)-independent subgroup of PKCs also become active when associated with PS at elevated level of DAG. Sustained activation of PKCs leads to the conversion of these enzymes into membrane-inserted and membrane protein-associated forms, which may be responsible for certain long-term neural responses. Activation of PKC results in the phosphorylation of cellular proteins; among them, several calmodulin (CaM)-binding proteins are the prominent substrates of these kinases. Phosphorylation of these proteins by PKC favors the release of CaM, which is required for the Ca2+/CaM-dependent enzymes. Thus, activation of PKCs can lead to diverse cellular responses through such amplification steps. Future studies should be directed at the elucidation of the activation of each PKC isoform in vivo to correlate with the physiological responses.

Developmental expression of protein kinase C isozymes in rat cerebellum.

Previously we showed that protein kinase C (PKC) isozymes (types I, II, and III) have distinctive neuronal localizations in cerebellum. In the present study, we followed the different appearances of these isozymes during the postnatal development of cerebellum. By immunoblot analysis, type I PKC was found to be low within 2 weeks after birth; an abrupt increase was observed between 2 and 3 weeks and leveled off afterwards. By immunofluorescent staining, the type I PKC-specific antibody recognized the cell bodies and dendrites of Purkinje cells. The increase of this isozyme between 2 and 3 weeks of age correlates with the spreading of Purkinje cell arborization, at which time bulk of synaptogenesis between dendritic spines and axons of granule cells occurs. Both type II and III PKCs were present in granule cells. At birth, the level of type II PKC was relatively high compared to that of type III PKC, and the type II PKC-specific antibody stained the granule cell precursors in the external layer more heavily than did the type III PKC-specific antibody. The level of type II PKC declined slightly after birth and increased again at one week and plateaued after three weeks, whereas that of type III PKC increased gradually until leveling off after three weeks. Throughout the development, the type III PKC-specific antibody also stained the cell bodies of Purkinje cells but not their dendrites. These results demonstrate that the developmental expression of PKC isozymes is under separate control, and their distinct cellular and subcellular localizations suggest their unique functions in the cerebellum.

Role of protein kinase C in cellular regulation.

Protein kinase C (PKC) consists of a family of closely related enzymes ubiquitously present in animal tissues. These enzymes respond to second messengers, Ca2+, diacylglycerol and arachidonic acid, to express their activities at membrane locations. Numerous hormones, neurotransmitters, growth factors and antigens are believed to transmit their signals by activation of a variety of phospholipases to generate these messengers. The various PKC isozymes, which exhibit distinct biochemical characteristics and unique cellular and subcellular localizations, may be differentially stimulated depending on the duration and strength of these messengers. Activation of PKC has been linked to the regulation of cell surface receptors, ion channels, secretion, gene expression, and neuronal plasticity and toxicity. The mechanisms of action of PKC in the regulation of these cellular functions are not entirely clear. Further study to identify the target substrates relevant to the various cellular functions is essential to define the functional diversity of this enzyme family.

Methylphenidate improves the behavioral and cognitive deficits of neurogranin knockout mice.

Neurogranin (Ng), a brain-specific calmodulin-binding protein, is expressed highly in hippocampus, and is important for cognitive function. Deletion of the Ng gene from mice caused attenuation of signal reaction cascade in hippocampus, impairments in learning and memory and high frequency stimulation-induced long-term potentiation (LTP). Environmental enrichment alone failed to improve cognitive function. In this study, behavioral testing revealed that Ng knockout (NgKO) mice were both hyperactive and socially withdrawn. Methylphenidate (MPH) was given to mice while they were also kept under an enrichment condition. MPH treatment reduced the hyperactivity of NgKO mice tested in both the open field and forced swim chamber. MPH improved their social abilities such that mice recognized and interacted better with novel subjects. The cognitive memories of MPH-treated mutants were improved in both water maze and contextual fear conditioning tests. High frequency stimulation-induced LTP of NgKO mice was also improved by MPH. The present treatment regimen, however, did not fully reverse the deficits of the mutant mice. In contrast, MPH exerted only a minimal effect on the wild type mice. At the cellular level, MPH increased the number of glial fibrillary acidic protein-positive cells in hippocampus, particularly within the dentate gyrus of NgKO mice. Therefore it will be of interest to determine the nature of MPH-mediated astrocyte activation and how it may modulate behavior in future studies. Taken together these NgKO mice may be useful for the development of better drug treatment to improve cognitive and behavioral impairments.

Stimulation-mediated translocation of calmodulin and neurogranin from soma to dendrites of mouse hippocampal CA1 pyramidal neurons.

Calmodulin (CaM) and neurogranin (Ng) are two abundant neuronal proteins in the forebrain whose interactions are implicated in the enhancement of synaptic plasticity. To gain further insight into the actions of these two proteins we investigated whether they co-localize in principle neurons and whether they respond to high frequency stimulation in a coordinated fashion. Immunohistochemical staining of CaM and Ng in mouse hippocampal slices revealed that CaM was highly concentrated in the nucleus of CA1 pyramidal neurons, whereas Ng was more broadly localized throughout the soma and dendrites. The asymmetrical localization of CaM in the nucleus of pyramidal neurons was in sharp contrast to the distribution observed in pyramidal cells of the neighboring subiculum, where CaM was uniformly localized throughout the soma and dendrites. The somatic concentrations of CaM and Ng in CA1 pyramidal neurons were approximately 10- and two-fold greater than observed in the dendrites, respectively. High frequency stimulation (HFS) of hippocampal slices promoted mobilization of CaM and Ng from soma to dendrites. These responses were spatially restricted to the area close to the site of stimulation and were inhibited by the N-methyl-D-asparate receptor antagonist 2-amino-5-phosphonopentanoic acid. Furthermore, HFS failed to promote translocation of CaM from soma to dendrites of slices from Ng knockout mice, which also exhibited deficits in HFS-induced long-term potentiation. Translocated CaM and Ng exhibited distinct puncta decorating the apical dendrites of pyramidal neurons and appeared to be concentrated in dendritic spines. These findings suggest that mobilization of CaM and Ng to stimulated dendritic spines may enhance synaptic efficacy by increasing and prolonging the Ca2+ transients and activation of Ca2+/CaM-dependent enzymes.