Regulation of gene expression by neurotransmitters in the central nervous system.
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Animals↗
Biomedical subjects
J L Martin
Publications and source records attributed to J L Martin.
Evidence supporting the existence of an activity-dependent astrocyte-neuron lactate shuttle.
Mounting evidence from in vitro experiments indicates that lactate is an efficient energy substrate for neurons and that it may significantly contribute to maintain synaptic transmission, particularly during periods of intense activity. Since lactate does not cross the blood-brain barrier easily, blood-borne lactate cannot be a significant source. In vitro studies by several laboratories indicate that astrocytes release large amounts of lactate. In 1994, we proposed a mechanism whereby lactate could be produced by astrocytes in an activity-dependent, glutamate-mediated manner. Over the last 2 years we have obtained further evidence supporting the notion that a transfer of lactate from astrocytes to neurons might indeed take place. In this article, we first review data showing the presence of mRNA encoding for two monocarboxylate transporters, MCT1 and MCT2, in the adult mouse brain. Second, by using monoclonal antibodies selectively directed against the two distinct lactate dehydrogenase isoforms, LDH1 and LDH5, a specific cellular distribution between neurons and astrocytes is revealed which suggests that a population of astrocytes is a lactate 'source' while neurons may be a lactate 'sink'. Third, we provide biochemical evidence that lactate is interchangeable with glucose to support oxidative metabolism in cortical neurons. This set of data is consistent with the existence of an activity-dependent astrocyte-neuron lactate shuttle for the supply of energy substrates to neurons.
Animals↗
The proposed Stark II regulations. What physicians should know.
Although the proposed regulations do not yet have the force and effect of law and HCFA is still seeking input in a variety of substantive areas, the regulations as proposed reflect HCFA's current view of enforcement of Stark II. Although simply complying with the statutory language and the Stark I regulations may be enough to avoid scrutiny until the proposed regulations are promulgated in final form, this approach may set physicians and physician groups up for significant disruption in their relationships in the future, particularly when proposed regulations have called certain practices into question. Clearly, every relationship between a physician (and his or her physician group) and any entity that provides DHS should be reexamined in light of the proposed regulations. In addition, arrangements under negotiation should be structured with the proposed new requirements in mind.
Conflict of Interest↗
Small heat shock proteins and protection against ischemic injury in cardiac myocytes.
BACKGROUND: Overexpression of the inducible hsp70 protects against ischemic cardiac damage. However, it is unclear whether the small heat shock proteins hsp27 and alphaB-crystallin protect against ischemic injury. METHODS AND RESULTS: Our aim was to examine whether the overexpression of hsp27 and alphaB-crystallin in neonatal and adult rat cardiomyocytes would protect against ischemic injury. Recombinant adenovirus expressing hsp27 or alphaB-crystallin under the control of the cytomegalovirus promoter was used to infect cardiac myocytes at high efficiency as assessed by immunostaining. Overexpression was confirmed by Western blot analysis. Cardiomyocytes were subjected to simulated ischemic stress, and survival was estimated through assessment of lactate dehydrogenase and creatine phosphokinase release. The hsp27 overexpression decreased lactate dehydrogenase release by 45+/-7.5% in adult cardiomyocytes but had no effect in the neonatal cells. In contrast, alphaB-crystallin overexpression was associated with a decrease in cytosolic enzyme release in both adult (29+/-6.6%) and neonatal (32+/-5.4%) cardiomyocytes. Decreased endogenous hsp25 with an antisense adenovirus produced a 29+/-9.9% increase in damage with simulated ischemia. Overexpression of the inducible hsp70 in adult cardiomyocytes was associated with a 34+/-4.6% decrease in lactate dehydrogenase release and is in line with our previous results in neonatal cardiomyocytes. CONCLUSIONS: The increased expression of hsp27 and alphaB-crystallin through an adenovirus vector system protects against ischemic injury in adult cardiomyocytes. Likewise, the overexpression of alphaB-crystallin protects against ischemic damage in neonatal cardiomyocytes. Decreasing the high levels of endogenous hsp25 present in neonatal cardiomyocytes renders them more susceptible to damage caused by simulated ischemia.
Adenoviridae↗
Comparison of lactate transport in astroglial cells and monocarboxylate transporter 1 (MCT 1) expressing Xenopus laevis oocytes. Expression of two different monocarboxylate transporters in astroglial cells and neurons.
The transport of lactate is an essential part of the concept of metabolic coupling between neurons and glia. Lactate transport in primary cultures of astroglial cells was shown to be mediated by a single saturable transport system with a Km value for lactate of 7.7 mM and a Vmax value of 250 nmol/(min x mg of protein). Transport was inhibited by a variety of monocarboxylates and by compounds known to inhibit monocarboxylate transport in other cell types, such as alpha-cyano-4-hydroxycinnamate and p-chloromercurbenzenesulfonate. Using reverse transcriptase-polymerase chain reaction and Northern blotting, the presence of mRNA coding for the monocarboxylate transporter 1 (MCT1) was demonstrated in primary cultures of astroglial cells. In contrast, neuron-rich primary cultures were found to contain the mRNA coding for the monocarboxylate transporter 2 (MCT2). MCT1 was cloned and expressed in Xenopus laevis oocytes. Comparison of lactate transport in MCT1 expressing oocytes with lactate transport in glial cells revealed that MCT1 can account for all characteristics of lactate transport in glial cells. These data provide further molecular support for the existence of a lactate shuttle between astrocytes and neurons.
Animals↗
Depletion of striatal dopamine transporter does not affect psychostimulant-induced locomotor activity.
The effect of neurotoxin-induced depletion of striatal dopamine transporter (DAT) binding sites on animals' responses to psychostimulants was investigated. Multiple 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or methamphetamine (METH) injections but not a single METH injection to Swiss Webster mice resulted in > 60% depletion of striatal DAT. MPTP-induced depletion of DAT did not affect METH- and cocaine-stimulated locomotor activity compared with the response of control mice. Pre-exposure to either the neurotoxic or the single non-neurotoxic dose of METH resulted in a marked locomotor sensitization in response to METH or cocaine challenge injections. The present results indicate that > 60% loss in striatal DAT binding sites has no effect on animals' responses to psychostimulants, and suggest that neural systems other than striatal DAT may contribute to the induction of locomotor sensitization to METH and cocaine.
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
Crystal structure at 1.1 A resolution of alpha-conotoxin PnIB: comparison with alpha-conotoxins PnIA and GI.
Conotoxins are small, cysteine-rich peptides isolated from the venom of Conus spp. of predatory marine snails, which selectively target specific receptors and ion channels critical to the functioning of the neuromuscular system. alpha-Conotoxins PnIA and PnIB are both 16-residue peptides (differing in sequence at only two positions) isolated from the molluscivorous snail Conus pennaceus. In contrast to the muscle-selective alpha-conotoxin GI from Conus geographus, PnIA and PnIB block the neuronal nicotinic acetylcholine receptor (nAChR). Here, we describe the crystal structure of PnIB, solved at a resolution of 1.1 A and phased using the Shake-and-Bake direct methods program. PnIB crystals are orthorhombic and belong to the space group P212121 with the following unit cell dimensions: a = 14.6 A, b = 26.1 A, and c = 29.2 A. The final refined structure of alpha-conotoxin PnIB includes all 16 residues plus 23 solvent molecules and has an overall R-factor of 14.7% (R-free of 15.9%). The crystal structures of the alpha-conotoxins PnIB and PnIA are solved from different crystal forms, with different solvent contents. Comparison of the structures reveals them to be very similar, showing that the unique backbone and disulfide architecture is not strongly influenced by crystal lattice constraints or solvent interactions. This finding supports the notion that this structural scaffold is a rigid support for the presentation of important functional groups. The structures of PnIB and PnIA differ in their shape and surface charge distribution from that of GI.
Amino Acid Sequence↗
Spectral equilibration and primary photochemistry in Heliobacillus mobilis at cryogenic temperature.
We performed multicolor femtosecond transient absorption measurements on membranes of the photosynthetic bacterium Heliobacillus mobilis at 20 K, by selective excitation at either the red or the blue extreme of the bacteriochlorophyll g Q(Y) band, which is split in three spectral forms (Bchl g 778, 793, and 808) at low temperature. In contrast to room temperature, there is no observable uphill energy transfer upon excitation at the red extreme. This provides a direct experimental confirmation of the expected strong temperature dependence of uphill energy transfer in multichromophore systems. Upon excitation at the blue edge, downhill energy transfer is observed on time ranges varying over 2 orders of magnitude and is discussed in terms of four distinct energy transfer processes: Bchl g 778* --> Bchl g 793* (approximately 50 fs); Bchl g 778* --> Bchl g 808* (approximately 400 fs); Bchl g 793* --> Bchl g 808* (approximately 1.4 ps); and within Bchl g 808* (approximately 7 ps). Surprisingly, the amount of oxidized primary donor P798+ formed on the time scale of picoseconds and tens of picoseconds was found to depend on the excitation conditions: trapping occurs mainly in approximately 80 ps and slower from directly excited Bchl g 808* and can additionally occur in a few picoseconds from Bchl g 778* and Bchl g 793* upon blue excitation. This finding implies that spectral equilibration is not complete prior to charge separation and furthermore is inconsistent with a funnel model, in which P798 is surrounded by long-wavelength pigments. More generally, we discuss to what extent our data bring constraints on the spatial distribution of the different spectral forms of the pigments.
Bacteria↗
Structure of TcpG, the DsbA protein folding catalyst from Vibrio cholerae.
The efficient and correct folding of bacterial disulfide bonded proteins in vivo is dependent upon a class of periplasmic oxidoreductase proteins called DsbA, after the Escherichia coli enzyme. In the pathogenic bacterium Vibrio cholerae, the DsbA homolog (TcpG) is responsible for the folding, maturation and secretion of virulence factors. Mutants in which the tcpg gene has been inactivated are avirulent; they no longer produce functional colonisation pili and they no longer secrete cholera toxin. TcpG is thus a suitable target for inhibitors that could counteract the virulence of this organism, thereby preventing the symptoms of cholera. The crystal structure of oxidized TcpG (refined at a resolution of 2.1 A) serves as a starting point for the rational design of such inhibitors. As expected, TcpG has the same fold as E. coli DsbA, with which it shares approximately 40% sequence identity. In addition, the characteristic surface features of DsbA are present in TcpG, supporting the notion that these features play a functional role. While the overall architecture of TcpG and DsbA is similar and the surface features are retained in TcpG, there are significant differences. For example, the kinked active site helix results from a three-residue loop in DsbA, but is caused by a proline in TcpG (making TcpG more similar to thioredoxin in this respect). Furthermore, the proposed peptide binding groove of TcpG is substantially shortened compared with that of DsbA due to a six-residue deletion. Also, the hydrophobic pocket of TcpG is more shallow and the acidic patch is much less extensive than that of E. coli DsbA. The identification of the structural and surface features that are retained or are divergent in TcpG provides a useful assessment of their functional importance in these protein folding catalysts and is an important prerequisite for the design of TcpG inhibitors.
Amino Acid Sequence↗
Crystal structure of Escherichia coli xanthine phosphoribosyltransferase.
Xanthine phosphoribosyltransferase (XPRT; EC 2.4.2.22) from Escherichia coli is a tetrameric enzyme having 152 residues per subunit. XPRT catalyzes the transfer of the phosphoribosyl group from 5-phospho-alpha-D-ribosyl 1-pyrophosphate (PRib-PP) to the 6-oxopurine bases guanine, xanthine, and hypoxanthine to form GMP, XMP, and IMP, respectively. Crystals grown in the absence of substrate or product were used to determine the structure of XPRT at a resolution of 1.8 A, by multiple isomorphous replacement. The core structure of XPRT includes a five-stranded parallel beta-sheet surrounded by three alpha-helices, which is similar to that observed in other known phosphoribosyltransferase (PRTase) structures. The XPRT structure also has several interesting features. A glutamine residue in the purine binding site may be responsible for the altered 6-oxopurine base specificity seen in this enzyme compared to other 6-oxopurine PRTases. Also, we observe both a magnesium ion and a sulfate ion bound at the PRib-PP binding site of XPRT. The sulfate ion interacts with Arg-37 which has a cis-peptide conformation, and the magnesium ion interacts with Asp-89, a highly conserved acidic residue in the PRib-PP binding site motif. The XPRT structure also incorporates a feature which has not been observed in other PRTase structures. The C-terminal 12 residues of XPRT adopt an unusual extended conformation and make interactions with a neighboring subunit. The very last residue, Arg-152, could form part of the active site of a symmetry-related subunit in the XPRT tetramer.
Amino Acid Sequence↗
The 27-kDa heat shock protein facilitates basic fibroblast growth factor release from endothelial cells.
Basic fibroblast growth factor is an important mitogenic and angiogenic factor that stimulates endothelial cell growth and migration. This hormone is not secreted via the classical vesicular pathway, and the identification of intracellular proteins that facilitate its release remains lacking. Transfection and expression of the 27-kDa human heat shock protein in bovine arterial endothelial cells doubles the rate of estrogen-induced basic fibroblast growth factor secretion, preferentially inducing the release of high molecular weight forms of the hormone. The secreted basic fibroblast growth factor is mitogenic to breast adenocarcinoma cells cultured in the conditioned medium obtained from the transfected endothelial cells. In contrast, decreasing the level of the endogenous heat shock protein homolog with an antisense vector markedly decreases basic fibroblast growth factor release. Anti-heat shock protein or anti-basic fibroblast growth factor antibodies co-precipitate both proteins from endothelial cell extracts, demonstrating a direct association between the two proteins. This interaction is likely to be an important step in the mechanism of basic fibroblast growth factor secretion.
Animals↗
Risk factors for adolescent pregnancy: how important is child sexual abuse? Otago Women's Health Study.
AIM: To examine the relative importance of child sexual abuse as a risk factor for adolescent pregnancy. METHOD: Postal survey and interview of a community sample of New Zealand women. RESULTS: Four preceding familial and psychosocial factors were associated with adolescent pregnancy; these included living in nonnuclear family or one in which the parents had frequent rows, being physically punished after the age of 12 and not having a confidante as a child. Whilst women who reported child sexual abuse were more likely to become pregnant under 19, this variable was confounded by these other characteristics. Only when child sexual abuse was of the most intrusive subtype, ie intercourse, did it independently predict adolescent pregnancy. Some interesting negative (nonsignificant) findings such as the type of high school attended, coeducational or single sex, were found. Material deprivation, as measured by parental socioeconomic and educational status, seemed less important in leading to adolescent pregnancy than has been reported from overseas. CONCLUSIONS: Adolescent pregnancy does not occur randomly in the community but was found in women who came from families with preexisting psychosocial problems. This suggests that preventive strategies aiming to reduce adolescent pregnancy should focus on measures which improve the general functioning of family units, in addition to providing good sexual information.
Adolescent↗
Regulation of energy metabolism by neurotransmitters in astrocytes in primary culture and in an immortalized cell line.
Evidence suggests that astrocytes might play an important role in cerebral energy metabolism. A recently developed cell line, called DI TNC1, displays several characteristic features of astrocytes. Thus, we have investigated in these cells a number of parameters related to energy metabolism. First, glycogen, the major energy reserve in the brain, is present in these cells and its levels are influenced by the glucose content of the growth medium and the presence of serum. Second, several neurotransmitters including noradrenaline and vasoactive intestinal peptide (VIP) induce a glycogenolytic response. Their effect on glycogen is paralleled by a similar effect on the formation of cyclic AMP, which is presumably the second messenger involved. Third, noradrenaline stimulates glucose utilization (as reflected by 2-deoxyglucose uptake) in DI TNC1 cells, an effect which is mimicked by the second messenger arachidonate. Interestingly, two actions of neurotransmitters, which are well characterized in primary astrocytes, are absent in DI TNC1 cells. These are the noradrenaline- and VIP-induced resynthesis of glycogen and the glutamate-stimulated glycolysis. In summary, the observations reported here lend further support to the concept that astrocytes are important for the control of brain energy metabolism. In addition, DI TNC1 cells might represent an interesting preparation to help decipher some of the astrocytic functions related to energy metabolism.
Animals↗
The uncharged surface features surrounding the active site of Escherichia coli DsbA are conserved and are implicated in peptide binding.
DsbA is a protein-folding catalyst from the periplasm of Escherichia coli that interacts with newly translocated polypeptide substrate and catalyzes the formation of disulfide bonds in these secreted proteins. The precise nature of the interaction between DsbA and unfolded substrate is not known. Here, we give a detailed analysis of the DsbA crystal structure, now refined to 1.7 A, and present a proposal for its interaction with peptide. The crystal structure of DsbA implies flexibility between the thioredoxin and helical domains that may be an important feature for the disulfide transfer reaction. A hinge point for domain motion is identified-the type IV beta-turn Phe 63-Met 64-Gly 65-Gly 66, which connects the two domains. Three unique features on the active site surface of the DsbA molecule-a groove, hydrophobic pocket, and hydrophobic patch-form an extensive uncharged surface surrounding the active-site disulfide. Residues that contribute to these surface features are shown to be generally conserved in eight DsbA homologues. Furthermore, the residues immediately surrounding the active-site disulfide are uncharged in all nine DsbA proteins. A model for DsbA-peptide interaction has been derived from the structure of a human thioredoxin:peptide complex. This shows that peptide could interact with DsbA in a manner similar to that with thioredoxin. The active-site disulfide and all three surrounding uncharged surface features of DsbA could, in principle, participate in the binding or stabilization of peptide.
Amino Acid Sequence↗
Structural analysis of three His32 mutants of DsbA: support for an electrostatic role of His32 in DsbA stability.
DsbA, a 21-kDa protein from Escherichia coli, is a potent oxidizing disulfide catalyst required for disulfide bond formation in secreted proteins. The active site of DsbA is similar to that of mammalian protein disulfide isomerases, and includes a reversible disulfide bond formed from cysteines separated by two residues (Cys30-Pro31-His32-Cys33). Unlike most protein disulfides, the active-site disulfide of DsbA is highly reactive and the oxidized form of DsbA is much less stable than the reduced form at physiological pH. His32, one of the two residues between the active-site cysteines, is critical to the oxidizing power of DsbA and to the relative instability of the protein in the oxidized form. Mutation of this single residue to tyrosine, serine, or leucine results in a significant increase in stability (of approximately 5-7 kcal/mol) of the oxidized His32 variants relative to the oxidized wild-type protein. Despite the dramatic changes in stability, the structures of all three oxidized DsbA His32 variants are very similar to the wild-type oxidized structure, including conservation of solvent atoms near the active-site residue, Cys30. These results show that the His32 residue does not exert a conformational effect on the structure of DsbA. The destabilizing effect of His32 on oxidized DsbA is therefore most likely electrostatic in nature.
Binding Sites↗
General anesthesia and elderly surgical patients.
As the elderly population continues to expand, more elderly patients will undergo surgical procedures. This patient population provides challenges for perioperative nurses, surgeons, and anesthesia care providers. By understanding normal age-related body system changes that may affect anesthesia, perioperative management team members can prevent complications and provide positive outcomes for elderly surgical patients.
Aged↗
Interpreting laboratory values in elderly surgical patients.
Although certain laboratory test results are expected to be outside the normal adult range in elderly patients, perioperative nurses must understand that these "normal abnormal" values are among the factors that make elderly patients more vulnerable to surgical risks. When preoperative testing reveals abnormalities, surgical procedures may be delayed while metabolic problems are treated. Prompt treatment of abnormal laboratory values and closer observation by perioperative nurses when elderly surgical patients laboratory values are borderline can help prevent negative consequences of surgical interventions in these patients.
Adult↗
Assessment and management of sensory loss in elderly patients.
The increasing number of elderly patients undergoing surgical procedures is a reality. Providing nursing care to these patients presents a challenge throughout their perioperative experiences. Knowledge of normal age-related changes coupled with data from preoperative assessments allows perioperative nurses to plan, implement, and evaluate care of elderly surgical patients with sensory losses.
Aged↗