Bedside multimarker testing for risk stratification in chest pain units: The chest pain evaluation by creatine kinase-MB, myoglobin, and troponin I (CHECKMATE) study.

BACKGROUND: Earlier, rapid evaluation in chest pain units may make patient care more efficient. A multimarker strategy (MMS) testing for several markers of myocardial necrosis with different time-to-positivity profiles also may offer clinical advantages. METHODS AND RESULTS: We prospectively compared bedside quantitative multimarker testing versus local laboratory results (LL) in 1005 patients in 6 chest pain units. Myoglobin, creatine kinase-MB, and troponin I were measured at 0, 3, 6, 9 to 12, and 16 to 24 hours after admission. Two MMS were defined: MMS-1 (all 3 markers) and MMS-2 (creatine kinase-MB and troponin I only). The primary assessment was to relate marker status with 30-day death or infarction. More patients were positive by 24 hours with MMS than with LL (MMS-1, 23.9%; MMS-2, 18.8%; LL, 8.8%; P=0.001, all comparisons), and they became positive sooner with MMS-1 (2.5 hours, P=0.023 versus LL) versus MMS-2 (2.8 hours, P=0.026 versus LL) or LL (3.4 hours). The relation between baseline MMS status and 30-day death or infarction was stronger (MMS-1: positive, 18.8% event rate versus negative, 3.0%, P=0.001; MMS-2: 21.9% versus 3.2%, P=0.001) than that for LL (13.6% versus 5.5%, P=0.038). MMS-1 discriminated 30-day death better (positive, 2.0% versus negative, 0.0%, P=0.007) than MMS-2 (positive, 1.8% versus negative, 0.2%; P=0.055) or LL (positive, 0.0% versus negative, 0.5%; P=1.000). CONCLUSIONS: Rapid multimarker analysis identifies positive patients earlier and provides better risk stratification for mortality than a local laboratory-based, single-marker approach.

Cellular mechanisms underlying carbachol-induced oscillations of calcium-dependent membrane current in smooth muscle cells from mouse anococcygeus.

1. At a holding potential of -40 mV, carbachol (50 microM) produced a complex pattern of inward currents in single smooth muscle cells freshly isolated from the mouse anococcygeus. Membrane currents were monitored by the whole-cell configuration of the patch-clamp technique. Previous work has identified the first, transient component as a calcium-activated chloride current (ICl(Ca)) and the second sustained component as a store depletion-operated non-selective cation current (I(DOC)). The object of the present study was to examine the cellular mechanisms underlying the third component, a series of inward current oscillations (I(oscil)) superimposed on I(DOC). 2. Carbachol-induced I(oscil) (amplitude 97 +/- 11 pA; frequency 0.26 +/- 0.02 Hz) was inhibited by the chloride channel blocker anthracene-9-carboxylic acid (A-9-C; 1 mM), and by inclusion of 1 mM EGTA in the patch-pipette filling solution. 3. In calcium-free extracellular medium (plus 1 mM EGTA), carbachol produced an initial burst of oscillatory current which lasted 94 s before decaying to zero; I(oscil) could be restored by re-admission of calcium. The frequency, but not the amplitude, of I(oscil) increased with increasing concentrations of extracellular calcium (0.5-10 mM). 4. Inclusion of the inositol triphosphate (IP3) receptor antagonist heparin (5 mg ml(-1) in the patch-pipette filling solution, or pretreatment of cells with the sarcoplasmic reticulum (SR) calcium ATPase inhibitor cyclopiazonic acid (CPA; 10 microM), prevented the activation of I(oscil) by carbachol. Caffeine (10 mM) activated both ICl(Ca) and I(DOC) and prevented the induction of I(oscil) by carbachol. Caffeine and CPA also abolished I(oscil) in the presence of carbachol, as did both a low (3 microM) and a high (30 microM) concentration of ryanodine. 5. Carbachol-induced I(oscil) was abolished by the general calcium entry blocker SKF 96365 (10 MM) and by Cd2+ (100 microM), but was unaffected by La3+ (400 microM). As found previously, I(DOC) was also blocked by SKF 96365 and Cd2+, but not La3+; the inhibition of I(DOC) preceded the abolition of I(oscil) by 27 s with SKF 96365 and by 30 s with Cd2+. Nifedipine (1 microM) produced a partial inhibition of the carbachol-induced I(oscil) frequency at holding potentials of -20 mV and -60 mV and, in addition, reduced I(DOC) at -60 mV by 18%. 6. It is concluded that carbachol-induced inward current oscillations in mouse anococcygeus cells are due to a calcium-activated chloride current, and reflect oscillatory changes in cytoplasmic calcium ion concentration. These calcium oscillations are derived primarily from the SR stores, but entry of calcium into the cell is necessary for store replenishment and maintenance of the oscillations. Capacitative calcium entry (via I(DOC) appears to be important not only for sustained contraction of this tissue, but also as a route for re-filling of the SR and, therefore, represents an important target for the development of novel and selective drugs.

Early diagnostic efficiency of cardiac troponin I and Troponin T for acute myocardial infarction.

OBJECTIVE: To compare the early diagnostic efficiency of the cardiac troponin I (cTn-I) level with that of the cardiac troponin T (cTn-T) level, as well as the creatine kinase (CK), CK-MB, and myoglobin levels, for acute myocardial infarction (AMI) in patients without an initially diagnostic ECG presenting to the ED within 24 hours of the onset of their symptoms. METHODS: A prospective, observational, cohort study was performed involving chest pain patients admitted to a large urban community hospital. Participants were consecutive consenting ED chest pain patients > or = 30 years of age. Exclusions included duration of symptoms > 24 hours, inability to complete data collection, receipt of CPR, and ST-segment elevation on the initial ECG. Measurements included levels of cTn-I, cTn-T, CK, CK-MB, and myoglobin at the time of presentation and 1, 2, 6, and 12-24 hours after presentation as well as presenting ECG and clinical follow-up. Confirmation of the diagnosis of AMI was based on World Health Organization criteria. RESULTS: Of the 177 patients included in the study, 27 (15%) were diagnosed as having AMIs. The sensitivities of all 5 biochemical markers for AMI were poor at the time of ED presentation (3.7-33.3%) but rose significantly over the study period. The sensitivity of cTn-T was significantly better than that of cTn-I over the initial 2 hours, but both markers' sensitivities were low (< 60%) during this time frame. The cTn-I was significantly more specific for AMI than was the cTn-T, but not significantly better than CK-MB or myoglobin. Likelihood ratio analysis showed that the biochemical markers with the highest positive likelihood ratios for AMI during the first 2 hours following ED presentation were myoglobin and CK-MB. From 6 through 24 hours, the positive likelihood ratios for cTn-I, CK-MB, and myoglobin were superior to those of CK and cTn-T. CONCLUSIONS: cTn-I, CK-MB, and myoglobin are significantly more specific for AMI than are CK and cTn-T. Myoglobin is the biochemical marker having the highest combination of sensitivity, specificity, and negative predictive value for AMI within 2 hours of ED presentation. Neither cTn-I nor cTn-T offers significant advantages over myoglobin and CK-MB in the early (< or = 2 hours) initial screening for AMI. The cardiac troponins are of benefit in identifying AMI > or = 6 hours after presentation.

Inhibition by sodium nitroprusside of a calcium store depletion-activated non-selective cation current in smooth muscle cells of the mouse anococcygeus.

1. The effects of sodium nitroprusside (SNP) on the non-selective cation current activated in response to intracellular calcium store depletion were studied using the whole-cell patch-clamp technique in single smooth muscle cells isolated from the mouse anococcygeus. Voltage-dependent calcium currents were blocked with extracellular nifedipine, and caesium and tetraethylammonium chloride were used to block voltage-dependent potassium currents. Calcium stores were depleted with caffeine (10 mM), carbachol (50 microM) or cyclopiazonic acid (CPA 10 microM; an inhibitor of the sarcoplasmic reticulum [SR] calcium-ATPase). 2. At a holding potential of -40 mV, both CPA and caffeine activated inward currents which consisted of two clearly distinguishable components; an initial transient current followed by a smaller sustained current. In the case of CPA, the amplitudes of the transient and sustained components were 19.7 +/- 2.1 pA and 3.5 +/- 0.3 pA respectively, whilst the equivalent values for caffeine were 188 +/- 21 and 4.8 +/- 0.3 pA. As described previously, the transient current results from activation of a calcium-dependent chloride conductance whilst the sustained current is a non-selective cation current, activated following intracellular calcium store depletion. 3. The muscarinic receptor agonist, carbachol, also activated a transient followed by a sustained current with amplitudes of 238 +/- 55 and 4.7 +/- 0.5 pA respectively. Superimposed on the sustained current were regular, oscillations of calcium-activated chloride current. 4. Both the transient and the sustained currents activated by CPA were absent in cells pretreated with SNP (10 microM). Application of SNP to a cell following activation of the sustained current by CPA inhibited the current by 88.6 +/- 3.8%. SNP (10 microM) did not inhibit the transient current activated by caffeine but abolished the sustained current. 5. SNP (10 microM) had no effect on the initial transient current activated by carbachol (50 microM). However, it did inhibit the oscillations in the inward current. In recordings from cells bathed in extracellular solution containing the chloride channel blocker, anthracene-9-carboxylic acid (A-9-C; 1 mM), carbachol activated only a sustained current. This current was inhibited by 88.1 +/- 6.5% by a concomitant application of SNP (10 microM) and was absent in cells pretreated with the nitrovasodilator. 6. The effects of SNP on the currents activated by caffeine (10 mM) were mimicked by 8-bromo-cyclic GMP (200 microM); thus the nucleotide had no effect on the transient current activated by caffeine but abolished the sustained current. The effects of SNP, but not those of 8-bromo-cyclic GMP, were inhibited by the nitric oxide-sensitive guanylyl cyclase inhibitor, 1H-[1, 2, 4]oxadiazolo[4, 3-a]quinoxaline-1-one (ODQ; 1 microM). ODQ alone produced a significant increase in the size of the sustained current activated by caffeine (7.8 +/- 0.7 pA). 7. These findings suggest that SNP activates guanylyl cyclase to inhibit the non-selective cation current activated as a result of intracellular calcium store depletion in mouse anococcygeus cells. Since the non-selective cation current appears to underlie the calcium entry process responsible for maintaining the sustained contractions to agonists in this tissue, this action of SNP may represent an important mechanism by which nitrates relax non-vascular smooth muscle.

Two distinct membrane currents activated by cyclopiazonic acid-induced calcium store depletion in single smooth muscle cells of the mouse anococcygeus.

1. By use of the whole-cell configuration of the patch-clamp technique, membrane currents induced by cyclopiazonic acid (CPA; an inhibitor of the sarcoplasmic reticulum (SR) calcium-ATPase) were investigated in single smooth muscle cells freshly dispersed from the mouse anococcygeus. Voltage-dependent calcium currents were blocked with extracellular nifedipine and caesium and tetraethylammonium chloride were used to block voltage-dependent potassium currents. 2. At a holding potential of -40 mV, CPA (10 microM) activated an inward current that consisted of two distinct components. The first was an initial transient current with an amplitude of 19.6 +/- 1.9 pA while the second was sustained and had an amplitude of 3.5 +/- 0.3 pA. 3. The current-voltage (I-V) relationship for the transient current showed marked outward rectification. The current had a reversal potential of 9.1 +/- 1.1 mV which was shifted to 29.0 +/- 4.2 mV when the extracellular chloride concentration was lowered from 148.4 to 58.4 mM. The sustained current had a near-linear I-V relationship and a reversal potential of 31.0 +/- 2.7 mV. Removal of extracellular calcium had no effect on the transient current, but shifted the reversal potential of the sustained current to 18.2 +/- 5.7 mV. 3. The initial transient current was abolished in cells bathed in extracellular solutions containing the chloride channel blockers, 4,4' diisothiocyanato-stilbene-2,2'-disulphonic acid (DIDS; 1 mM) or anthracene-9-carboxylic acid (A-9-C; 1 mM), and was absent in cells containing the calcium buffers EGTA (1 to 5 mM) or BAPTA (10 mM). The second sustained current was unaffected by either the chloride channel blockers or the intracellular calcium buffers. 4. Treatment of the cells with caffeine (10 mM) produced similar inward currents to those produced by CPA. In the presence of caffeine, CPA (10 microM) induced no further inward current. 5. In organ bath studies, CPA (10 microM)-induced contractions of the mouse anococcygeus were inhibited by cadmium and nickel (both 50-400 microM) and the general calcium entry blocker, SKF 96365 (10 microM); lanthanum and gadolinium had no effect at concentrations up to 400 microM. The pharmacology of the CPA-induced non-selective cation current mirrored that of the CPA-induced whole muscle contraction being reversed by cadmium (100 microM) and SKF 96365 (10 microM), but unaffected by lanthanum (400 microM). The initial chloride conductance was unaffected by cadmium, SKF 96365 or lanthanum. 6. It is concluded that CPA activates a transient calcium-dependent chloride current as a consequence of calcium release from intracellular stores; this current would result in depolarization and opening of voltage-operated calcium channels, which mediate the nifedipine-sensitive component of muscle contraction. In addition, as a result of emptying the SR, CPA activates a non-selective cation conductance which may underlie the nifedipine-insensitive calcium entry process utilised during sustained contraction.

The nitrergic transmitter of the anococcygeus--NO or not?

Nonadrenergic noncholinergic (NANC) relaxations of the anococcygeus muscle are reduced by inhibitors of nitric oxide synthase (NOS). Since NOS can be detected within 6-hydroxydodpamine-resistant nerve tracts running through the muscle, it seems clear that these NANC relaxations result from activation of the L-arginine/NO pathway within the prejunctional nerve terminal, an example of so-called "nitrergic" transmission. However, a number of substances (hydroquinone, superoxide anions, hydroxocobalamin) profoundly reduce relaxations to exogenous NO but do not affect those to nitrergic field stimulation; such observations have raised questions over the nature of the substance actually released from the nitrergic nerves. Several possible explanations are discussed: (1) NO is released attached to a carrier molecule, perhaps in the form of a nitrosothiol; (2) NO is released in a modified redox form; (3) NO is released as a free radical, but is protected within the neuroeffector junction by other substances which preferentially interact with scavenger molecules; and (4) NO is released as a free radical and, because of a rapid and unhindered rate of diffusion over short distances (100-200 microM), it is less susceptible than exogenous NO to scavenger molecules. As yet, there is insufficient experimental evidence to decide which, if any, of these explanations is correct.

Variable potency of nitrergic-nitrovasodilator relaxations of the mouse anococcygeus against different forms of induced tone.

1. U46619 (thromboxane A2 receptors; 0.002-1 microM), carbachol (muscarinic M3 receptors; 0.1-100 microM), cyclopiazonic acid (CPA; Ca(2+)-ATPase inhibitor; 0.1-30 microM) and K+ (5-100 mM) produced concentration-dependent contractions of the mouse isolated anococcygeus muscle. Equi-effective, submaximal concentrations of each agent were used in further experiments (40 nM U46619; 5 microM carbachol; 5 microM CPA; 70 mM K+). 2. Nifedipine (1 microM) totally abolished contractile responses to K+; those to U46619, carbachol and CPA were reduced by only 20-30% in the presence of nifedipine, but were greatly reduced (> 90%) by a combination of nifedipine and SKF 96365 (0.1-40 microM). 3. In Ca(2+)-free medium, contractions to K+ and CPA were abolished. Small residual responses remained to both carbachol and U46619; those to carbachol were transient, could not be repeated in the continued absence of Ca2+ and were prevented by pre-incubation with CPA, but unaffected by SKF 96365; those to U46619 were sustained, could be repeated in the absence of Ca2+, and were resistant to CPA and SKF 96365. 4. Tone induced by all four agents could be relaxed by sodium nitroprusside (SNP), but with a clear order of potency. SNP (pIC40) was most effective against U46619 (7.92), less so against carbachol (6.80) and CPA (6.68), and least potent against K+ (5.94). A similar order of potency was observed with 8Br-cyclic GMP (50 microM) and nitrergic field stimulation (1-20 Hz). 5. The relaxant potency of SNP was similar in normal Krebs solution and in high K+ (70 mM) Krebs containing 1 microM nifedipine. 6. Inclusion of SNP (0.01-1 microM) or 8Br-cyclic GMP (50 microM) in the Ca2+-free medium inhibited the transient residual response to carbachol. Inclusion of similar concentrations of SNP or 8Br-cyclic GMP,during Ca2+ re-loading, increased the subsequent residual contraction to carbachol in Ca2+-free medium.7. At higher concentrations, SNP (0.1-10 microM) produced a partial relaxation of the sustained contraction to U46619 in Ca2+-free medium.8. Thus, the relaxant potency of the nitrergic stimuli was dependent on the agent and mechanism used to induce tone in the preparation. Examination of the contractile/relaxant interactions suggests that altered Ca2+ sequestration and inhibition of contractile protein function may underlie nitrergic relaxations of this tissue.

Value of serial myoglobin levels in the early diagnosis of patients admitted for acute myocardial infarction.

STUDY OBJECTIVE: The potential for missing the diagnosis of acute myocardial infarction (AMI) and the need for appropriate use of ICU beds make early and accurate diagnostic tests to assist in this diagnosis valuable. We studied the use of serial myoglobin determinations for patients evaluated in the emergency department and admitted for possible AMI. DESIGN: Over a 3.5-month period, all patients presenting to the ED and admitted for suspected cardiac symptoms had serial cardiac enzymes obtained prospectively at admission and 2, 3, 4, and 6 hours after the onset of symptoms. SETTING: Large urban community hospital. PARTICIPANTS: One hundred thirty-three consecutive patients admitted to treat or rule out AMI. RESULTS: Twenty-one of 22 patients with an initially normal myoglobin that doubled within 1 to 2 hours after presentation were positive for AMI (specificity, 95%). Sensitivity of myoglobin at 2 hours after the onset of symptoms was 37% and rose to 86% at 6 hours, with 95% specificity. The negative predictive value if myoglobin was normal at 6 hours and had not doubled within 2 hours was 97% (positive predictive value, 88%). CONCLUSION: A repeat myoglobin level that doubled within 1 to 2 hours after the initial value, even if still within the normal range, was highly specific for AMI. Serial myoglobin levels may be useful in earlier identification of AMI to help prevent inappropriate discharge from the ED and for appropriate placement in ICU beds.

Nitrergic stimulation does not inhibit carbachol-induced inositol phosphate generation in the rat anococcygeus.

Carbachol (50 microM) produced a rapid, transient increase in inositol-1,4,5-trisphosphate (IP3) levels in the rat anococcygeus; the peak increase observed at 10 s (3-fold above controls) was greatly reduced in the presence of atropine (100 nM), but was unaffected by nitrergic stimulation (10 Hz), sodium nitroprusside (10 microM) or 8-Br-cyclic GMP (200 microM). Following loading of muscles with [3H]myo-inositol, subsequent exposure to carbachol for 30 min resulted in a 6-fold increase in the accumulation of [3H]inositol-1-monophosphate; again, this action of carbachol was greatly attenuated by atropine, but unaffected by nitrergic stimulation, sodium nitroprusside or 8-Br-cyclic GMP. It is concluded that inhibition of agonist-induced generation of inositol phosphates cannot explain the ability of nitrergic activation to relax (by 54-62%) carbachol-induced tone in this tissue.

Localisation of nitric oxide synthase within non-adrenergic, non-cholinergic nerves in the mouse anococcygeus.

Immunocytochemical staining of whole mount preparations of the mouse anococcygeus muscle, using antibodies to rat brain nitric oxide synthase (NOS), revealed a dense network of NOS-immunoreactive nerve fibres running through the tissue. These fibres were resistant to the sympathetic neurotoxin 6-hydroxydopamine and are therefore likely to be the non-adrenergic nerves which mediate relaxation of this smooth muscle. Further, NOS-immunoreactive fibres were absent following denervation by cold-storage (4 degrees C; 72 h), which has been shown to abolish non-adrenergic, non-cholinergic (NANC) relaxations. The results provide strong support for the hypothesis that the L-arginine:NO pathway is responsible for the generation of the NANC transmitter in the anococcygeus.

The influence of L-NG-nitro-arginine on sympathetic nerve induced contraction and noradrenaline release in the rat isolated anococcygeus muscle.

1. The possibility of an interaction between the motor sympathetic and inhibitory non-adrenergic, non-cholinergic (NANC) nerves in the rat anococcygeus was investigated using L-NG-nitro-arginine (L-NOARG), an inhibitor of L-arginine: NO synthase. 2. L-NOARG (50 microM) increased contractions induced by field stimulation (20 s trains; 0.5-40 Hz); overall, the frequency-response curve was displaced six-fold to the left. D-NOARG (50 microM) was without effect. 3. The potentiation produced by L-NOARG was reversed by 200 microM L-, but not D-, arginine. 4. L-NOARG had no effect on contractions induced by exogenous noradrenaline (NA) or on field stimulation-induced overflow of tritium from muscles previously loaded with [3H]-NA. 5. It is concluded that the endogenous nitrate NANC transmitter does not influence release of NA from the sympathetic nerves and the potentiation of contractions induced by field stimulation in the presence of L-NOARG most probably results from removal of the opposing relaxing influence of concomitantly released NANC transmitter.

An investigation of some S-nitrosothiols, and of hydroxy-arginine, on the mouse anococcygeus.

1. The effect of five S-nitrosothiols, and of the stereoisomers of NG-hydroxy-arginine (HOARG), were investigated on the mouse anococcygeus. 2. All five S-nitrosothiols produced concentration-related (0.1-100 microM) relaxations of carbachol (50 microM)-induced tone; the order of potency was S-nitroso-L-cysteine (CYSNO) > S-nitroso-N-acetyl-D,L-penicillamine (SNAP) > S-nitrosoglutathione (GSNO) > S-nitrosocoenzyme A (CoASNO) > S-nitroso-N-acetyl-L-cysteine (NACNO). The relaxations were unaffected by the nitric oxide synthase (NOS) inhibitor, L-NG-nitro-arginine (10 microM) (L-NOARG). 3. Cold-storage of the tissue for 72 h resulted in loss of sympathetic and non-adrenergic, non-cholinergic (NANC) nerve function. NOS activity in the tissue was reduced by 97%. Despite this, relaxations induced by the S-nitrosothiols were unaffected. 4. Haemoglobin (50 microM) attenuated relaxations induced by NO and the S-nitrosothiols, although responses to 3-isobutyl-1-methyl-xanthine were unaffected. N-methyl-hydroxylamine (2 mM) which has been shown previously to produce selective inhibition of NANC and nitrovasodilator responses in this tissue, also reduced responses to all S-nitrosothiols. 5. Hydroquinone (100 microM) greatly reduced relaxations to CYSNO (by 88%) but had no effect on those to SNAP, GSNO, CoASNO or NACNO. Since hydroquinone does not reduce responses to NANC stimulation, CYSNO is unlikely to be the NANC transmitter. 6. L-HOARG by itself (up to 100 microM) had no significant effect on carbachol-induced tone or on NANC (10 Hz; 10 strain every 100 s) relaxations. However, it produced reversal of the inhibitory effects of L-NOARG (10;pM), being only slightly less potent than L-arginine. D-HOARG was without effect.L-HOARG had no effect on relaxations induced by 1.51iM NO.7. The results show that S-nitrosothiols are potent relaxants of the mouse anococcygeus; they act directly on the smooth muscle with a mechanism similar to NO and other nitrovasodilators. In addition,the results are consistent with L-HOARG being an intermediate in the biosynthesis of NO from L-arginine, although there is no evidence for it acting to stabilize NO extracellularly.

Differentiation by hydroquinone of relaxations induced by exogenous and endogenous nitrates in non-vascular smooth muscle: role of superoxide anions.

1. The influence of hydroquinone on relaxations induced by nitric oxide (NO), nitrovasodilator drugs, and non-adrenergic, non-cholinergic (NANC) field stimulation has been investigated in three tissues in which endogenous nitrates have been implicated in the NANC response; the mechanism of action of hydroquinone was also studied. 2. In mouse anococcygeus, hydroquinone (10-100 microM) produced a concentration-dependent inhibition of relaxations induced by 15 microM NO. Hydroquinone, 100 microM, which reduced responses to NO by 85%, had no effect on relaxations induced by NANC field stimulation (10 Hz; 20s trains), hydroxylamine (10 microM), sodium nitroprusside (1 microM) or sodium azide (20 microM). 3. In guinea-pig trachea, 100 microM hydroquinone reduced relaxations to 150 microM NO by 75%, but had no effect on those to NANC stimulation (10 Hz; 30 s trains) or sodium azide (5 microM). 4. In rat gastric fundus, 100 microM hydroquinone reduced relaxations to 1 microM NO by 85%, but had no effect on those to NANC stimulation (0.5 Hz; 15 s trains) or sodium azide (2 microM). 5. Superoxide dismutase (SOD; 50 u ml-1) had no effect on relaxations of the mouse anococcygeus in response to 15 microM NO or 10 Hz NANC stimulation. Further, the inhibition of responses to NO by hydroquinone was unaffected in the presence of SOD. 6. Hydroquinone (10-100 microM) failed to generate superoxide anions, as detected by a chemiluminescent assay. However, 100 microM hydroquinone, like SOD (50 u ml-1), produced almost complete inhibition of superoxide anion chemiluminescence induced by xanthine (500 microM): xanthine oxidase (0.07 u ml-1). 7. It is concluded that, in our system, hydroquinone inhibits NO by acting as a free radical scavenger rather than by generating superoxide anions. The ability of hydroquinone to block relaxations to NO, but not NANC stimulation, may suggest that the endogenous nitrate substance released by these NANC nerves may not be free NO, but may be an NO-containing, or NO-generating, molecule.

The influence of L-NG-nitro-arginine on field stimulation induced contractions and acetylcholine release in guinea pig isolated tracheal smooth muscle.

The interaction between parasympathetic and inhibitory non-adrenergic, non-cholinergic nerves in tracheal smooth muscle was investigated by determining the effects of the NO-synthase inhibitor L-NG-nitro-arginine (L-NOARG) on contractions and the associated acetylcholine release elicited by field stimulation of the muscle. At frequencies above 2Hz contractile responses to field stimulation were potentiated by L-NOARG (50 microM). alpha-chymotrypsin pre-treatment potentiated contractile responses at all frequencies, but the effects of L-NOARG were unaltered. The effect of L-NOARG on responses to 5Hz electrical stimulation was not mimicked by D-NOARG, was reversed by L-, but not D-arginine and was unaffected by epithelium removal. L-NOARG did not affect responses to exogenous acetylcholine nor the overflow of 3H from tissues previously loaded with [3H]-choline. It is therefore concluded that field stimulation of tracheal smooth muscle induces the release of an endogenous nitrate, which, by an inhibitory action on smooth muscle, functionally antagonises the concomitantly released parasympathetic neurotransmitter.

Cyclic nucleotide content of the rat anococcygeus during relaxations induced by drugs or by non-adrenergic, non-cholinergic field stimulation.

Low concentrations of sodium nitroprusside (0.2 and 1 microM) relaxed carbachol-induced tone of the rat anococcygeus but did not affect the content of either cGMP or cAMP; higher concentrations (10,100 and 1000 microM) produced greater relaxation (greater than 60%) and a rise in cGMP but not cAMP. In the presence of the cGMP-phosphodiesterase inhibitor M&B 22948 (10 microM), 1 microM sodium nitroprusside produced greater relaxation and a selective increase in cGMP. Forskolin (0.5-250 microM) caused relaxation and a selective increase in cAMP; the concentration-response relationships of the two effects were similar. Non-adrenergic, non-cholinergic (NANC) field stimulation (10 Hz; 20 s trains) reduced tone by 52% but had no effect on cyclic nucleotide content; in the presence of 10 microM M&B 22948 or 1 microM sodium nitroprusside, NANC stimulation produced a greater degree of relaxation and increased cGMP but not cAMP content. The results show that NANC stimulation acts like sodium nitroprusside, causing a selective increase in cGMP, and this supports the proposal that NANC transmission in the rat anococcygeus involves an endogenous nitrate; the possibility that multiple pools of cGMP exist in the anococcygeus is discussed.

L-NG-nitro arginine inhibits non-adrenergic, non-cholinergic relaxations of guinea-pig isolated tracheal smooth muscle.

The effects of L-NG-nitro arginine (L-NOARG) on alpha-chymotrypsin-resistant, non-adrenergic, non-cholinergic (NANC) relaxations of guinea-pig tracheal smooth muscle have been examined. L-NOARG (1-100 microM), but not D-NOARG (100 microM), inhibited the NANC relaxations in a concentration-related manner. The effects of L-NOARG were partially reversed by L-arginine but not D-arginine. L-NOARG was without effect on acetylcholine-induced contractile responses of the trachea or on relaxations produced by vasoactive intestinal peptide, sodium nitroprusside or isoprenaline. These results suggest that an endogenous nitrate may contribute to NANC relaxations of tracheal smooth muscle.

Inhibition of colonization of the chicken alimentary tract with Salmonella typhimurium gram-negative facultatively anaerobic bacteria.

Oral administration of strains of food poisoning salmonellas to day-old chickens produced a profound inhibition in the subsequent colonization of the caeca by a strain of Salmonella typhimurium given one day later. Closely related genera were unable to produce a similar inhibition. The inhibition was not the result of bacteriophages produced by the first strain. Neither was it the result of an immunological response by the host induced by the first strain. In additional experiments in day-old chickens, inhibition of an Escherichia coli Nalr strain and of a Citrobacter sp. Nalr strain was produced by the antibiotic-sensitive forms of the homologous strains while strains from other genera did not produce any inhibition. When an avirulent mutant of S. typhimurium was used for pre-treatment a statistically significant reduction in the excretion of the super-infecting S. typhimurium Nalr strain over several weeks was produced. A genus specific inhibition was reproduced in vitro by mixed culture experiments. Live cultures were necessary for in vitro inhibition. Killed cells or a culture supernatant produced no inhibition.

Calcium: limited indications, some danger.

Calcium chloride has been advocated since the 1920s for the resuscitation of asystole, electromechanical dissociation (EMD), and ventricular fibrillation. Reports of side effects and complications have been numerous. Studies of calcium assays following American Heart Association recommended dosages have shown dangerously elevated serum levels. Large retrospective clinical studies in Milwaukee and Tampa have found no evidence of improved survival with calcium chloride in asystole and EMD. A prospective randomized double-blind study comparing calcium chloride and saline controls in the Milwaukee Paramedic system for asystole and EMD using standard AHA protocols showed no statistically significant difference in resuscitation rates or long-term survival between the calcium and no-calcium groups for the rhythm of asystole. Although patients with EMD had statistically improved resuscitation rates when calcium chloride was given, only one of the patients survived to hospital discharge. Because of the low rates of resuscitation and long-term survival in patients presenting in asystole and EMD, proving that calcium chloride does not enhance survival would require large multicenter trials. However, since no controlled study has ever documented significant benefit, its routine use in asystole and EMD cannot be supported. Calcium has long been used in medical treatment of hypocalcemic and hyperkalemic states and should be administered in moribund patients who have the proper clinical history and clinical signs of hypocalcemia.

Inhibition of colonization of the chicken caecum with Salmonella typhimurium by pre-treatment with strains of Escherichia coli.

Simultaneous oral administration of broth cultures of three strains of Escherichia coli isolated from sewage and an abattoir strongly inhibited the colonization of a subsequently administered strain of Salmonella typhimurium. The three strains were protective against the S. typhimurium strain under a variety of conditions: in different breeds and in chickens fed different diets. The strains were not equally effective against other salmonella strains. Oral administration of the strains produced a statistically significant reduction in the excretion of the S. typhimurium strain over a period of 7 weeks.

Decision making in prehospital sudden cardiac arrest.

Many studies of prehospital resuscitation report on selected populations. We examined a series of 445 unselected nontraumatic cardiac arrests. Emergency cardiac care (ECC) was not initiated in 126 (28%). ECC was begun in 319 (78%), but was terminated in 132 (33%). Ninety-four (21%) were admitted to the hospital with palpable pulses and organized rhythm (successful resuscitation/save rate for patients presenting in ventricular fibrillation was 50%/25%. Multivariate regression analysis was used to identify the relative importance of significant variables in predicting survival, and the analysis identified the presence of ventricular fibrillation, short paramedic response times, and short paramedic treatment times.