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Vasodilator flare due to activation of superficial cutaneous afferents in humans: heat-sensitive versus histamine-sensitive fibers.

Histamine-sensitive nerve endings are assumed to terminate in the superficial epidermis. Heat-sensitive nociceptors that are excited by brief carbondioxide-laser pulses must also terminate within the epidermis, because this infrared radiation has an extinction length of about 10 microns. We now compared laser heat stimuli (10 W, 50 ms, 20 mm2) with intradermal injections of histamine (10(-10) to 10(-8) mol) in their capacity to cause cutaneous vasodilatation (flare) in awake human subjects. Cutaneous blood flow was measured with a two-channel laser-Doppler device. The radiant heat pulses caused a transient cutaneous vasodilatation that spread at least 15 mm from the stimulus site. There was no visible flare and vasodilatation could only be detected by laser Doppler measurements. Although the heat pulses elicited enough nociceptor activity to be perceived as moderately painful, the magnitude of the vasodilatation was smaller and its duration shorter than after the smallest dose of histamine. In contrast, nociceptor activation by heat is usually stronger than by histamine. These data indicate that flare and pain are two different aspects of cutaneous small fiber function.

Histamine↗

Role of ATP-sensitive K+ channels in anoxia-sensitive negative potential of endolymph.

To investigate the possible role of the ATP-sensitive K+ channels (KATP channels) in the generation of the anoxia-sensitive negative potential (ASNP), the effects of the treatment with glybenclamide and diazoxide on the endocochlear potential (EP) and K+ activity (AK) in the scala media were examined with double-barrelled K(+)-selective microelectrodes. The experiments were carried out in guinea pig cochleae, using glybenclamide as a KATP channel blocker and diazoxide as a KATP channel opener. Perilymphatic perfusion of glybenclamide decreased the amplitude of the ASNP and shortened its duration, whereas perfusion of diazoxide increased ASNP amplitude and prolonged its duration. Glybenclamide enhanced the decrease of endolymphatic AK by anoxia, whereas diazoxide suppressed this AK decrease. The results suggest that KATP channels may be involved in the generation of the ASNP.

Adenosine Triphosphate↗

Mouse and rat strain variations in sensitivity to N-nitroso-diethylamine, hereditary transmission of the trait and the effect of 3-tert-butyl-4-hydroxyanisole on sensitivity.

1. Strain variations among male mice were studied in terms of the number of days of survival with chronic administration of N-nitroso-diethylamine (NDEA). Four inbred strains, two F1 progenies and one F2 progeny were tested. 2. BALB/c mice survived for the longest period, whereas C3H mice survived for the shortest time. Results of examinations of BALB/c-C3H-F1, -F2 and C57BL-CBA-F1 mice revealed that the hereditary trait could be adequately explained by postulating two loci of genes or gene clusters that regulate the sensitivity to NDEA. 3. Simultaneous chronic administration of 3-tert-butyl-4-hydroxyanisole (BHA) could prolong the survival period. 4. Preliminary histopathological examinations of the liver tissues revealed that the lesion at the time of death of the mice varied considerably depending on the strain and the length of survival. Evidence for hereditary transmission of the characteristics of histopathological changes, including development of liver hemangiosarcoma, is presented. 5. The strain variations among male and female rats were also studied in terms of the number of days of survival with chronic administration of NDEA. Five strains and one F1 progeny were tested. 6. From these and previous observations, the possible biochemical factors determining sensitivity to NDEA were discussed.

Animals↗

Thymidine kinase deficient human cells have increased UV sensitivity in their capacity to support herpes simplex virus but normal UV sensitivity for colony formation.

A thymidine kinase deficient (tk-) and two thymidine kinase proficient (tk+) human cell lines were compared for UV sensitivity using colony-forming ability as well as their capacity to support the plaque formation of herpes simplex type 1 (HSV-1). The tk- line (143 cells) was a derivative of one of the tk+ lines (R970-5), whereas the other tk+ line (AC4 cells) was a derivative of the 143 cells obtained by transfection with purified sheared HSV-2 DNA encoding the viral tk gene. 143, R970-5 and AC4 cells showed a similar UV sensitivity for colony-forming ability. In contrast, the capacity to support HSV-1 plaque formation immediately (within 1 h) after UV-irradiation was reduced to a greater extent in the 143 cells compared to the R970-5 and AC4 cells. Capacity curves for plaque formation of the HSV-1: KOS wild-type (tk+) strain were similar to those for the HSV-1: PTK3B mutant (tk-) strain in the 3 cell strains, indicating that the viral tk gene does not influence the ability of HSV-1 to form plaques in UV-irradiated compared to unirradiated human cells. Cellular capacity for HSV-1 plaque formation was found to recover in both tk- and tk+ cells for cultures infected 24 h after UV-irradiation. These results suggest that repair of UV-damaged DNA takes place to a similar extent in both tk- and tk+ human cells, but the kinetics of repair are initially slower in tk- compared to tk+ human cells.

Animals↗

NK sensitivity of neuroblastoma cells determined by a highly sensitive coupled luminescent method.

The measurement of natural killer (NK) cells toxicity against tumor or virus-infected cells especially in cases with small blood samples requires highly sensitive methods. Here, a coupled luminescent method (CLM) based on glyceraldehyde-3-phosphate dehydrogenase release from injured target cells was used to evaluate the cytotoxicity of interleukin-2 activated NK cells against neuroblastoma cell lines. In contrast to most other methods, CLM does not require the pretreatment of target cells with labeling substances which could be toxic or radioactive. The effective killing of tumor cells was achieved by low effector/target ratios ranging from 0.5:1 to 4:1. CLM provides highly sensitive, safe, and fast procedure for measurement of NK cell activity with small blood samples such as those obtained from pediatric patients.

Cell Line, Tumor↗

Protein-sensitive hypoglycemia without leucine sensitivity in hyperinsulinism caused by K(ATP) channel mutations.

OBJECTIVE: Because children with congenital hyperinsulinism (HI) caused by recessive loss of function mutations in the adenosine triphosphate (ATP)-dependent potassium channel (K(ATP)-HI) are not leucine sensitive, we evaluated for protein-induced hypoglycemia with oral protein tolerance tests. STUDY DESIGN: Blood glucose and insulin concentrations were measured every 15 minutes for 3 hours after an oral protein load in children with K(ATP)-HI (n = 11) and compared with those of children with glutamate dehydrogenase HI (n = 12) and control subjects (n = 12). RESULTS: Similar to children with glutamate dehydrogenase HI, patients with K(ATP)-HI displayed protein-induced hypoglycemia (10/11) with blood glucose concentrations declining by 17 to 69 mg/dL. In contrast, oral protein had little effect on blood glucose concentrations in control subjects. CONCLUSIONS: Protein-induced hypoglycemia is a feature of K(ATP)-HI, despite the absence of leucine sensitivity. The results indicate that amino acids can stimulate insulin secretion via a glutamate dehydrogenase- and K(ATP) channel-independent pathway.

ATP-Binding Cassette Transporters↗

The effect of dibutyryl cAMP on tetrodotoxin-sensitive and -resistant voltage-gated sodium currents in rat dorsal root ganglion neurons and the consequences for their sensitivity to lidocaine.

Tetrodotoxin-sensitive (TTXS) sodium currents in dorsal root ganglia (DRG) neurons were enhanced by DcAMP applied acutely or by pre-treatment. Pre-treatment increased peak TTXS by 28%. This compared to the increase of tetrodotoxin-resistant sodium currents (TTXR) of 123%. In both cases the increase was associated with a hyperpolarizing shift in activation potentials. Slow inactivation was slower for both TTXR and TTXS in DcAMP treated neurons but rates of recovery from inactivation were not altered. Lidocaine blocked TTX-R with an IC(50) of 0.51+/-0.15mM (n=9) which was reduced to 0.14+/-0.05mM (n=8, P<0.05) in DcAMP treated cells. The sensitivity of TTX-S currents to lidocaine was not altered by DcAMP (control EC(50)=0.89+/-0.16mM, n=9; DcAMP EC(50)=0.73+/-0.19mM, n=6). It is concluded that TTXS currents in DRG are, like TTX-R currents, enhanced by cAMP but whilst the pharmacology of TTXR channels with respect to lidocaine is altered, that to TTXS channels is not.

Algorithms↗

Endothelin-1 inhibition of cardiac ATP-sensitive K+ channels via pertussis-toxin-sensitive G-proteins.

OBJECTIVE: Secretion of endothelin-1 (ET-1) and activation of cardiac ATP-sensitive K+ (KATP) channels are facilitated under myocardial metabolic stress. The aim of this study was to investigate the effects of ET-1 on KATP channels and to assess underlying mechanisms in ventricular myocytes. METHODS: Single channel currents were measured with the voltage-clamp technique in cell-attached patches from enzymatically-isolated single guinea pig ventricular myocytes. In some experiments, the open-cell-attached mode was employed by permeating the membrane with streptolysin-O. RESULTS: ET-1 concentration-dependently inhibited single KATP channel currents, which had been activated by metabolic poisoning, with an IC50 of 3.8 +/- 0.7 pM. BQ-123, an ETA receptor-selective antagonist, reduced the effects of ET-1. ET-1 effects were largely abolished in the myocytes pre-incubated with pertussis toxin. In the open-cell-attached mode, where the intracellular ATP concentration ([ATP]) could be virtually controlled, the effects of ET-1 were abolished. Muscarinic receptor stimulation inhibited the channels in a similar manner to ET-1, whereas beta-adrenoceptor stimulation accelerated channel activation. By analogy, ouabain also inhibited KATP channel activity under metabolic stress presumably because inhibition of the Na+/K+ pump spares subsarcolemmal ATP. ET-1 inhibited the KATP channels that had been reactivated in the continuous presence of ouabain. CONCLUSIONS: ET-1 reversibly inhibited KATP channels. This effect appears to be mediated by an increase in subsarcolemmal [ATP] which results from inhibition of adenylate cyclase activities through PTX-sensitive G-proteins coupled to ETA receptors.

Adenosine Triphosphate↗

Increased behavioral neurosteroid sensitivity in a rat line selectively bred for high alcohol sensitivity.

Acute administration of a neurosteroid 5beta-pregnan-3alpha-ol-20-one induced a greater impairment in motor performance of the selectively bred alcohol-sensitive (ANT) than alcohol-insensitive (AT) rats. This difference was not associated with the sensitivity of gamma-aminobutyrate type A (GABA(A)) receptors, as 5alpha-pregnan-3alpha-ol-20-one (allopregnanolone) decreased the autoradiographic signals of t-butylbicyclophosphoro[35S]thionate binding to GABA(A) receptor-associated ionophores more in the brain sections of AT than ANT rats. Nor was the difference associated with baseline levels of neuroactive progesterone metabolites, as 5alpha-pregnan-3,20-dione (5alpha-DHP) and 5alpha-pregnan-3alpha-ol-20-one were lower in the ANT rats. After ethanol (2 g/kg, i.p.) administration and the subsequent motor performance test, the increased brain concentrations of these metabolites were still lower in the ANT than AT rats, although especially in the cerebellum the relative increases were greater in the ANT than AT rats. The present data suggest that the mechanisms mediating neurosteroid-induced motor impairment are susceptible to genetic variation in rat lines selected for differences in ethanol intoxication.

Animals↗

Levosimendan, a novel Ca2+ sensitizer, activates the glibenclamide-sensitive K+ channel in rat arterial myocytes.

The electrophysiological effect of levosimendan, a novel Ca(2+)-sensitizing positive inotropic agent and vasodilator, was examined on rat mesenteric arterial myocytes using the patch clamp technique. Resting potential was significantly hyperpolarized with levosimendan, with an EC50 of 2.9 microM and maximal effect (19.5 +/- 3.5 mV; n = 12) at 10 microM. Levosimendan (10 microM) significantly increased the whole-cell outward current. The currents intersected close to the calculated EK (-84 mV), suggesting that the activated current was a K+ current. Hyperpolarization and stimulation of K+ current by levosimendan were not prevented by 30 microM H-7 (a non-specific inhibitor of protein kinases) and 100 nM charybdotoxin (a blocker of Ca(2+)-activated K+ channels), but were abolished by 10 microM glibenclamide. In single-channel current recording in open cell-attached patches, two types of K+ channels were observed having conductances of 26 and 154 pS. The 154 pS channels were not affected by levosimendan and glibenclamide. The 26 pS channels were evoked in one-fourth of the patches when 10 microM levosimendan (and 0.1 mM UDP) was added (at -60 mV) and channel activity was abolished by glibenclamide. The mean open probability of the 26 pS channels was 0.094 +/- 0.017 (n = 9), and the mean open time (at -60 mV) was 6.6 ms in the presence of UDP and levosimendan. Although significant hyperpolarization (4.7 +/- 1.5 mV, n = 8) was observed at 1 microM levosimendan, the same concentration did not affect Ca2+ channel currents (n = 10). In summary, levosimendan hyperpolarized the arterial myocytes, probably through activation of a glibenclamide-sensitive K+ channel. This mechanism may contribute to the vasodilating action of levosimendan.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

Effect of age on short-wavelength sensitive cone electroretinogram and long- and middle-wavelength sensitive cone electroretinogram.

Using a light-emitting diode (LED) built-in contact lens electrode, we studied the effect of age on short-wavelength sensitive cone electroretinograms (S-cone ERG) and long- and middle-wavelength sensitive cone ERGs (LM-cone ERG). We recorded ERGs in 31 pseudophakic subjects to avoid the effect of yellowing in the human crystalline lens. The intensities of our stimuli were on the asymptote of the intensity-response curve. Linear regression analysis against age was carried out on the S-cone ERG b-wave, and the LM-cone ERG a-, b-, and d-waves. We found significant age-dependent reduction in the amplitude of the S-cone ERG b-wave, and the LM-cone ERG a- and d-waves, significant prolongation in the peak time of the S-cone b-wave and the LM-cone ERG b-wave, significant increase in the b/a ratio of the LM-cone ERG, but no significant age correlation between the LM-cone ERG a-wave and the amplitude of the b-wave. Our results provide evidence that age-related changes in S- and LM-cone systems begin in early adulthood in the human eye. A significant increase in the b/a ratio suggests that off-bipolar cells are more vulnerable to aging than on-bipolar cells.

Adult↗

Blockade of ATP-sensitive K+ channels attenuates preconditioning effect on myocardial metabolism in swine: myocardial metabolism and ATP-sensitive K+ channels.

OBJECTIVE: We investigated if blockade of ATP-sensitive K+ channels (KATP) abolishes the protective effect of ischemic preconditioning (IP) on myocardial metabolism and ischemia-induced reactive hyperemia (RH) in pigs. METHODS: IP was elicited by a single cycle of 5 min occlusion and 5 min reperfusion of coronary artery, followed by 15 min of test ischemia and 120 min of reperfusion. Vehicle or the ATP-sensitive K+ channels (KATP) blocker, glibenclamide (3 or 6 mg/kg; G3 or G6) was administered before IP (groups; IP, G3+IP, G6+IP). As respective controls, the same treatment was performed in groups without IP (groups; C, G3, G6). Tissue levels of ATP, creatine phosphate (CP) and intracellular pH (pHi) in the area at risk were measured by 31P-nuclear magnetic resonance spectroscopy. RH after 5 min of preconditioning ischemia was assessed by regional myocardial blood flow. RESULTS: ATP and pHi were preserved after 15 min of ischemia in the IP group [C/IP; ATP=57+/-4/76+/-10% of baseline, pHi=6.18+/-0.08/6.66+/-0.03, P<0.05, C vs. IP]. Both doses of glibenclamide completely abolished the ATP sparing effect of IP. The high dose completely abolished pHi preservation (G6+IP=6.33+/-0.06), while the low dose showed only a partial effect (G3+IP=6.48+/-0.03). Glibenclamide did not adversely affect myocardial metabolism in groups without IP. Glibenclamide attenuated RH after 5 min of ischemia by 30% in both subendocardium and subepicardium. CONCLUSIONS: Blockade of KATP abolished the preconditioning effect on myocardial metabolism, and partially attenuated post-ischemic reactive hyperemia in pigs. These results indicate that KATP activation might be involved in the mechanisms of these phenomena, reactive hyperemia is not sufficient to induce IP protection.

Adenosine Triphosphate↗

Induction of suppressor T cells to dinitrofluorobenzene contact sensitivity by application of sensitizer through Langerhans cell-deficient skin. II. Kinetics of the induction of suppressor T cells.

Tolerance to dinitrofluorobenzene (DNFB) contact sensitivity (DNFB CS) appeared in C3H/HeN mice within 2 days after application of DNFB on Langerhans cell-deficient tail skin. A single painting on tail skin led to complete prevention of DNFB CS for at least 14 days. Spleen cells from tolerant mice could transfer the suppression to syngeneic recipients, while the cells treated in vitro with anti-Thy 1.2 lost their suppressive activities completely. From these facts, this tolerance might be due to active suppression caused by suppressor T cells. These suppressor T cells were antigen-specific, because recipients exhibited normal CS to picryl chloride. Furthermore, the suppressor T cells were demonstrated to be afferent blockers in nature and their precursors were sensitive to cyclophosphamide. Therefore, the suppressor T cells induced by DNFB percutaneously administered through epidermal Langerhans cell-deficient region were thought to be generated in the same or in a very similar suppressor pathway as the intravenously administered dinitrobenzene sulfonate (DNBSO3), a soluble reactive dinitrophenyl congener.

Animals↗

The sensitivity to DNA single strand breakage in mitochondria, but not in nuclei, of Chinese hamster V79 and variant cells correlates with their cellular sensitivity to hydrogen peroxide.

To investigate whether a relation exists between the level of DNA damage by and cytotoxicity of hydrogen peroxide, we measured the initial level of H2O2-induced nuclear and mitochondrial DNA single strand breaks in Chinese hamster V79 and H2O2-resistant variant cells (Hpr-4) with an alkaline elution technique and a quantitative Southern blot technique, respectively. The frequency of DNA single strand breaks in mitochondrial DNA induced by H2O2 was more than one hundred times that of nuclear DNA in the parent V79 cells. While a similar frequency of nuclear DNA single strand breaks was generated in V79 and Hpr-4 cells at an equidose of H2O2, a lower number of mitochondrial DNA single strand breaks were generated in Hpr-4 cells than in V79 cells by H2O2 in the range of 100 microM to 5 mM. The sensitivity to mitochondrial DNA single strand break-induction correlated with the cellular sensitivity to H2O2 in Chinese hamster V79 and variant cells.

Alkalies↗

Taste reactivity in high-alcohol-sensitive and low-alcohol-sensitive rats.

High, low, and control alcohol-sensitive (HAS, LAS, CAS, respectively) rats were tested for their perception of the taste of alcohol using the taste reactivity test. Reactivity tests with a single concentration of sucrose and quinine were also done. After initial taste reactivity, all rats were tested for alcohol consumption in a standard two-bottle test (water in the second bottle). Postconsumption taste reactivity tests completed the experiment. Results indicated that HAS, LAS, and CAS rats did not differ significantly in their taste reactivity response to a range of alcohol concentrations (5-40%), nor did they differ significantly in response to sucrose or quinine. Reactivity responses were similar for each group before and after the consumption tests. Despite the lack of line differences in taste reactivity, HAS and LAS rats consumed significantly less alcohol than the CAS rats during the two-bottle access tests. The present results are in contrast to research done with rats selectively bred for alcohol consumption (Alcohol Preferring and Nonpreferring rats, High Alcohol Drinking and Low Alcohol Drinking rats), which exhibit clear line differences in patterns of reactivity changes following alcohol access. The selection phenotype of alcohol sensitivity appears to be independent of rats' behavioral response to the taste of alcohol.

Alcohol Drinking↗

Neonatal alcohol exposure produces hyperactivity in high-alcohol-sensitive but not in low-alcohol-sensitive rats.

Children of women who consume high amounts of alcohol during their pregnancies vary greatly in physical and behavioral outcomes. Although many factors, such as dose and timing of exposure, undoubtedly contribute to this variation, one important determinant may be genetic differences in the response to alcohol. The present study examined activity levels in high alcohol sensitivity (HAS) and low alcohol sensitivity (LAS) rats following neonatal alcohol exposure. These lines were selectively bred for extremes in ethanol-induced "sleep times." The HAS and LAS offspring were exposed to alcohol via an artificial rearing procedure using the "pup-in-the-cup" technique. Rat pups were exposed to ethanol (6 g/kg/day) from postnatal day (PD) 4 through 7 and faded to a dose of 3 g/kg/day on PD 8 and 9. An artificially reared gastrostomy control group (GC) and a normally reared suckle control group (SC) were also included. Activity level was measured on PD 18 through PD 21 for 30 min daily in automated activity monitors. Neonatal ethanol exposure produced overactivity in HAS rats, relative to their controls, but the same ethanol treatment had no effect on the LAS rats. Importantly, there were no differences in blood alcohol concentrations (around 420 mg/dl) between the two lines during the treatment period. These data suggest that genetic differences in response to alcohol may be a predictor for some of the behavioral teratogenic effects of alcohol.

Animals↗

Neonatal alcohol exposure produces more severe motor coordination deficits in high alcohol sensitive rats compared to low alcohol sensitive rats.

Prenatal exposure to alcohol can produce a number of behavioral alterations, including hyperactivity, learning deficits and motor impairments. However, the severity and nature of behavioral alterations varies markedly among children of women who drink during pregnancy. One important determinant of this variation may be genetic differences in the response to alcohol. Recently, we demonstrated that exposure to alcohol during development produced hyperactivity in rats bred for high alcohol sensitivity (HAS), but not in rats bred for low alcohol sensitivity (LAS). These lines were selectively bred for extremes in alcohol-induced "sleep time." The present study investigated the effects of ethanol exposure during development on motor coordination later in life in both HAS and LAS rats. Using an artificial rearing procedure, neonatal pups from each line were exposed to a binge-like alcohol treatment on postnatal days (PD) 4-9. Within each line, one group was exposed to ethanol (6.0 g/kg/day), one group served as an artificially reared control, and a third served as a normally reared control group. On PD 30, parallel bar motor performance was evaluated. Exposure to ethanol during development severely impaired motor performance in the HAS rats compared to their controls. In LAS rats, early ethanol exposure produced only mild and nonsignificant effects on motor performance. Thus, HAS rats were more vulnerable to ethanol-induced motor deficits compared to the LAS rats. Importantly, there were no differences in peak blood alcohol level between the lines, indicating that vulnerability to ethanol's teratogenic effects was not due to differences in metabolic rate. These results suggest that genetic differences in response to alcohol may serve as a predictor for susceptibility to ethanol's teratogenic effects.

Alcohol Drinking↗

Both insulin sensitivity and glucose sensitivity are impaired in patients with non-diabetic liver cirrhosis.

Background: Careful nutritional support is required in patients with liver cirrhosis due to their glucose intolerance. To elucidate the mechanism of glucose intolerance in cirrhotics, we measured insulin secretion, whole body insulin sensitivity (SI), and glucose sensitivity (SG) in non-diabetic cirrhotics.Methods: Eight patients with compensated cirrhosis who showed normal fasting blood glucose levels and non-diabetic curves on a 75 g oral glucose tolerance test participated in this study. Four normal volunteers were selected as controls. After an overnight fast, glucose was injected intravenously at 300 mg kg(-1) in 2 min followed 20 min later by intravenous insulin at 0.02 U kg(-1) in 5 min. Sequential blood samples were drawn from 20 min before the glucose injection to 3 h post-injection, and plasma glucose and insulin levels were determined. Plasma glucose and insulin disappearance curves were analyzed using the minimal compartment model, and kinetic parameters, including glucose clearance (KG), insulin secretion, SI and SG, were estimated.Results: KG was slower in cirrhosis than in controls, although not significant (P=0.051). Insulin secretion was not different between the two groups. However, SI was significantly lower in cirrhotics (0.814x10(-4) min(-1) pM(-1); 0.572-1.403x10(-4) min(-1) pM(-1)) as compared to controls (1.643x10(-4) min(-1) pM(-1); 0.678-2.085x10(-4) min(-1) pM(-1)) (P=0.029). SG was also lower in the cirrhosis (0.0154 min(-1); 0.0071-0.0208 min(-1)) than in the control group (0.0211 min(-1); 0.0184-0.0260 min(-1)) (P=0.026).Conclusion: Both SI and SG are already impaired in non-diabetic cirrhotic patients even when KG is minimally delayed and insulin secretion has not yet been affected.

Journal Article↗