Solution structure and excitation energy transfer in phycobiliproteins of Acaryochloris marina investigated by small angle scattering.

The structure of phycobiliproteins of the cyanobacterium Acaryochloris marina was investigated in buffer solution at physiological temperatures, i.e. under the same conditions applied in spectroscopic experiments, using small angle neutron scattering. The scattering data of intact phycobiliproteins in buffer solution containing phosphate can be well described using a cylindrical shape with a length of about 225Å and a diameter of approximately 100Å. This finding is qualitatively consistent with earlier electron microscopy studies reporting a rod-like shape of the phycobiliproteins with a length of about 250 (M. Chen et al., FEBS Letters 583, 2009, 2535) or 300Å (J. Marquart et al., FEBS Letters 410, 1997, 428). In contrast, phycobiliproteins dissolved in buffer lacking phosphate revealed a splitting of the rods into cylindrical subunits with a height of 28Å only, but also a pronounced sample aggregation. Complementary small angle neutron and X-ray scattering experiments on phycocyanin suggest that the cylindrical subunits may represent either trimeric phycocyanin or trimeric allophycocyanin. Our findings are in agreement with the assumption that a phycobiliprotein rod with a total height of about 225Å can accommodate seven trimeric phycocyanin subunits and one trimeric allophycocyanin subunit, each of which having a height of about 28Å. The structural information obtained by small angle neutron and X-ray scattering can be used to interpret variations in the low-energy region of the 4.5K absorption spectra of phycobiliproteins dissolved in buffer solutions containing and lacking phosphate, respectively.

Evaluation of the effect of fluconazole on the pharmacokinetics of cyclosporin A in healthy dogs after a single dose and at steady-state.

The aim of the study was to describe the effect of fluconazole on the pharmacokinetics of cyclosporin A in healthy dogs when investigated as a single dose and at steady-state. Five healthy adult dogs were used in the study in a crossover design receiving either 5 mg/kg of cyclosporin A (CsA) alone or 5 mg/kg of fluconazole with 2.5 mg/kg of cyclosporin A (CsA/Flu) for 35 days. Pharmacokinetic curves were performed on day 1 and day 35 in addition to sampling trough and suspected peak concentrations (C2) twice weekly with LC/MS/MS. There was no statistically significant difference noted in any pharmacokinetic value (AUC0-inf. [day 1, P = 0.225], AUCtau [day 35, P = 0.225], t½ [day 1, P = 0.279; day 35, P = 0.686], and Cmax [day 1, P = 0.225; day 35, P = 0.225]) between the treatment groups by sampling day. There was a statistically significant increase in AUC (CsA P = 0.043; CsA/Flu P = 0.043) and t½ (CsA P = 0.042, CsA/Flu P = 0.042) over time within each group. There were no significant differences in the Cmax (CsA P = 0.08; CsA/Flu P = 0.08) when comparing day 1 vs. day 35. Steady-state cyclosporine concentrations were achieved by day 10 in both groups. Subjectively, individual variability was noted among the dogs and a much larger sample size would be beneficial in a future study.

Excitation energy transfer and electron-vibrational coupling in phycobiliproteins of the cyanobacterium Acaryochloris marina investigated by site-selective spectroscopy.

In adaption to its specific environmental conditions, the cyanobacterium Acaryochloris marina developed two different types of light-harvesting complexes: chlorophyll-d-containing membrane-intrinsic complexes and phycocyanobilin (PCB) - containing phycobiliprotein (PBP) complexes. The latter complexes are believed to form a rod-shaped structure comprising three homo-hexamers of phycocyanin (PC), one hetero-hexamer of phycocyanin and allophycocyanin (APC) and probably a linker protein connecting the PBPs to the reaction centre. Excitation energy transfer and electron-vibrational coupling in PBPs have been investigated by selectively excited fluorescence spectra. The data reveal a rich spectral substructure with a total of five low-energy electronic states with fluorescence bands at 635nm, 645nm, 654nm, 659nm and a terminal emitter at about 673 nm. The electronic states at ~635 and 645 nm are tentatively attributed to PC and APC, respectively, while an apparent heterogeneity among PC subunits may also play a role. The other fluorescence bands may be associated with three different isoforms of the linker protein. Furthermore, a large number of vibrational features can be identified for each electronic state with intense phonon sidebands peaking at about 31 to 37cm⁻¹, which are among the highest phonon frequencies observed for photosynthetic antenna complexes. The corresponding Huang-Rhys factors S fall in the range between 0.98 (terminal emitter), 1.15 (APC), and 1.42 (PC). Two characteristic vibronic lines at about 1580 and 1634cm⁻¹ appear to reflect CNH⁺ and CC stretching modes of the PCB chromophore, respectively. The exact phonon and vibrational frequencies vary with electronic state implying that the respective PCB chromophores are bound to different protein environments. This article is part of a special issue entitled: photosynthesis research for sustainability: keys to produce clean energy.

Peripheral and site-specific CD4(+) CD28(null) T cells from rheumatoid arthritis patients show distinct characteristics.

Proinflammatory CD4(+) CD28(null) T cells are frequently found in the circulation of patients with rheumatoid arthritis (RA), but are less common in the rheumatic joint. In the present study, we sought to identify functional differences between CD4(+) CD28(null) T cells from blood and synovial fluid in comparison with conventional CD28-expressing CD4(+) T cells. Forty-four patients with RA, displaying a distinct CD4(+) CD28(null) T cell population in blood, were recruited for this study; the methylation status of the IFNG locus was examined in isolated T cell subsets, and intracellular cytokine production (IFN-γ, TNF, IL-17) and chemokine receptor expression (CXCR3, CCR6 and CCR7) were assessed by flow cytometry on T cells from the two compartments. Circulating CD4(+) CD28(null) T cells were significantly more hypomethylated in the CNS-1 region of the IFNG locus than conventional CD4(+) CD28(+) T cells and produced higher levels of both IFN-γ and TNF after TCR cross-linking. CD4(+) CD28(null) T cells from the site of inflammation expressed significantly more CXCR3 and CCR6 compared to their counterparts in blood. While IL-17A production could hardly be detected in CD4(+) CD28(null) cells from the blood, a significant production was observed in CD4(+) CD28(null) T cells from synovial fluid. CD4(+) CD28(null) T cells were not only found to differ from conventional CD4(+) CD28(+) T cells in the circulation, but we could also demonstrate that synovial CD4(+) CD28(null) T cells showed additional effector functions (IL-17 coproduction) as compared to the same subset in peripheral blood, suggesting an active role for these cells in the perpetuation of inflammation in the subset of patients having a CD28(null) population.

"Invisible" Detergents Enable a Reliable Determination of Solution Structures of Native Photosystems by Small-Angle Neutron Scattering.

Photosystems I (PSI) and II (PSII) are pigment-protein complexes capable of performing the light-induced charge separation necessary to convert solar energy into a biochemically storable form, an essential step in photosynthesis. Small-angle neutron scattering (SANS) is unique in providing structural information on PSI and PSII in solution under nearly physiological conditions without the need for crystallization or temperature decrease. We show that the reliability of the solution structure critically depends on proper contrast matching of the detergent belt surrounding the protein. Especially, specifically deuterated ("invisible") detergents are shown to be properly matched out in SANS experiments by a direct, quantitative comparison with conventional matching strategies. In contrast, protonated detergents necessarily exhibit incomplete matching so that related SANS results systematically overestimate the size of the membrane protein under study. While the solution structures obtained are close to corresponding high-resolution structures, we show that temperature and solution state lead to individual structural differences compared with high-resolution structures. We attribute these differences to the presence of a manifold of conformational substates accessible by protein dynamics under physiological conditions.

Current limits of structural biology: The transient interaction between cytochrome c 6 and photosystem I.

Trimeric photosystem I from the cyanobacterium Thermosynechococcus elongatus (TePSI) is an intrinsic membrane protein, which converts solar energy into electrical energy by oxidizing the soluble redox mediator cytochrome c 6 (Cyt c 6 ) and reducing ferredoxin. Here, we use cryo-electron microscopy and small angle neutron scattering (SANS) to characterize the transient binding of Cyt c 6 to TePSI. The structure of TePSI cross-linked to Cyt c 6 was solved at a resolution of 2.9 Å and shows additional cofactors as well as side chain density for 84% of the peptide chain of subunit PsaK, revealing a hydrophobic, membrane intrinsic loop that enables binding of associated proteins. Due to the poor binding specificity, Cyt c 6 could not be localized with certainty in our cryo-EM analysis. SANS measurements confirm that Cyt c 6 does not bind to TePSI at protein concentrations comparable to those for cross-linking. However, SANS data indicate a complex formation between TePSI and the non-native mitochondrial cytochrome from horse heart (Cyt c HH ). Our study pinpoints the difficulty of identifying very small binding partners (less than 5% of the overall size) in EM structures when binding affinities are poor. We relate our results to well resolved co-structures with known binding affinities and recommend confirmatory methods for complexes with K M values higher than 20 µM.

Excited state dynamics in recombinant water-soluble chlorophyll proteins (WSCP) from cauliflower investigated by transient fluorescence spectroscopy.

The present study describes the fluorescence emission properties of recombinant water-soluble chlorophyll (Chl) protein (WSCP) complexes reconstituted with either Chl a or Chl b alone (Chl a only or Chl b only WSCP, respectively) or mixtures of both pigments at different stoichiometrical ratios. Detailed investigations were performed with time and space correlated ps fluorescence spectroscopy within the temperature range from 10 to 295 K. The following points were found: (a) The emission spectra at room temperature (295 K) are well characterized by bands with a dominating Lorentzian profile broadened due to phonon scattering and peak positions located at 677, 684 and 693 nm in the case of Chl a only WSCP and at 665, 675 and 689 nm for Chl b only WSCP. In addition, all spectra contain minor bands in the longer wavelength region. (b) The emission spectra at 10 K of samples suspended in buffer containing 50% glycerol are dominated by bands peaking at 668 nm for Chl b only WSCP and at 685 nm for Chl a only WSCP and samples reconstituted with mixtures of Chl a and Chl b. (c) At 10 K and in buffer with 50% glycerol the decay kinetics of WSCP samples with Chl a only are dominated by a component with a time constant of 6.2 (+/-0.2) ns at 685 nm while those of WSCP containing mixtures of Chl a and Chl b are characterized by a slightly shorter value of 6.0 (+/-0.2) ns. WSCP containing Chl b only exhibits a distinctly longer value of 7.0 (+/-0.3) ns at an emission wavelength of 668 nm. (d) The decay associated emission spectra at 10 K of all samples exhibit at least 3 decay components with time constants of 80-120 ps, 2-4 ns and 6-7 ns in 50% glycerol. These results are consistently described within the framework of our previously presented model (J. Phys. Chem. B 2007, 111, No. 46, 13325; J. Phys. Chem. B 2007, 111, No. 35, 10487) , for the structural motifs of chlorophyll binding to the tetrameric protein matrix of WSCP. It is shown that formation of strongly coupled open sandwich dimers does not lead to quenching of 1Chl a* or 1Chl b*.

New sources and instrumentation for neutrons in biology.

Neutron radiation offers significant advantages for the study of biological molecular structure and dynamics. A broad and significant effort towards instrumental and methodological development to facilitate biology experiments at neutron sources worldwide is reviewed.

Brain stimulation and morphine reward deficits in dopamine D2 receptor-deficient mice.

RATIONALE: The rewarding effects of lateral hypothalamic brain stimulation, various natural rewards, and several drugs of abuse are attenuated by D1 or D2 dopamine receptor (D1R or D2R) antagonists. Much of the evidence for dopaminergic involvement in rewards is based on pharmacological agents with limited or "relative" selectivity for dopamine receptor subtypes. Genetically engineered animal models provide a complementary approach to pharmacological investigations. OBJECTIVES: In the present study, we explored the contribution of dopamine D2Rs to (1) brain stimulation reward (BSR) and (2) the potentiation of this behavior by morphine and amphetamine using D2R-deficient mice. METHODS: Wild-type (D2Rwt), heterozygous (D2Rhet), and D2R knockout (D2Rko) mice were trained to turn a wheel for rewarding brain stimulation. Once equivalent rate-frequency curves were established, morphine-induced (0, 1.0, 3.0, and 5.6 mg/kg s.c.) and amphetamine-induced (0, 1.0, 2.0, and 4.0 mg/kg i.p.) potentiations of BSR were determined. RESULTS: The D2Rko mice required approximately 50% more stimulation than the D2Rwt mice did. With the equi-rewarding levels of stimulation current, amphetamine potentiated BSR equally across the three genotypes. In contrast, morphine potentiated rewarding stimulation in the D2Rwt, had no effect in the D2Rhet, and antagonized rewarding stimulation in the D2Rko mice. CONCLUSIONS: D2R elimination decreases, but does not eliminate, the rewarding effects of lateral hypothalamic stimulation. After compensation for this deficit, amphetamine continues to potentiate BSR, while morphine does not.

Dose-response of inhaled diltiazem on airway reactivity to methacholine and exercise in subjects with mild asthma.

Methacholine challenges were performed by 10 asthmatic subjects, 2 hours before and 15 minutes after placebo (diluent alone) and 5, 10, 15, 30, and 60 mg inhaled diltiazem given in a single-blind crossover manner. There was no significant change from placebo in the dose of methacholine required to produce a 20% decrease in forced expiratory volume in the first second (FEV1) (PD20); the fold increase in PD20 from baseline was 1.1 +/- 0.1 after placebo, 1.4 +/- 0.2 after 5 mg, 1.8 +/- 0.3 after 10 mg, 1.4 +/- 0.2 after 15 mg, 1.6 +/- 0.2 after 30 mg, and 1.2 +/- 0.1 after 60 mg. There was a 1% chance that we missed a twofold difference between placebo and the 10 mg dose because of inadequate sample size. Fifteen minutes before a standardized exercise challenge, 10 subjects received placebo, 10 mg, and the highest dose tolerated during the methacholine study (20 to 45 mg) in a randomized double-blind crossover design. The mean +/- SE maximum postexercise decrease in FEV1 was 28.8% +/- 5.7% after placebo, 23.4% +/- 4.6% after 10 mg, and 20.8% +/- 3.0% after high-dose diltiazem (P greater than 0.05). There was a 12% chance that we missed a 15% difference between placebo and the high-dose regimen because of inadequate sample size. We conclude that diltiazem does not attenuate airway reactivity to methacholine or exercise even when high concentrations are delivered to the lungs.

Effects of diltiazem, dipyridamole, and their combination on hemostasis.

Calcium channel blockers and antiplatelet agents, alone and in combination, have been reported to induce bleeding in patients undergoing surgery. Since diltiazem and dipyridamole influence platelet function in vitro and in vivo, their influence on hemostasis was examined in five normal men given diltiazem, 90 mg by mouth, followed by 60 mg every 6 hr for 48 hr, or dipyridamole, 75 mg by mouth every 8 hr for 48 hr. At 24 hr, the alternate drug was added to the regimen to assess effects of the combination on hemostasis. Platelet aggregation, serum thromboxane B2 and 6-keto-PGF1 alpha concentrations (stable metabolites of thromboxane A2 and prostacyclin), bleeding time, prothrombin time, partial thromboplastin time, and serum diltiazem concentrations were measured. Diltiazem and dipyridamole alone and in combination had no significant effect on bleeding time, prothrombin time, or partial thromboplastin time. Platelet aggregation induced by threshold concentrations of adenosine diphosphate, epinephrine, and calcium ionophore A 23187 were inhibited by diltiazem and dipyridamole alone and in combination. The only change in prostaglandin concentrations was a slight increase in serum 6-keto-PGF1 alpha after diltiazem. Despite influences on platelet function, neither diltiazem nor dipyridamole alone or in combination induced clinically relevant changes in hemostasis.

Effect of calcium channel blockers on theophylline disposition.

Twelve healthy subjects received a single oral dose of theophylline, 5 mg/kg alone, and after 7 days of oral verapamil, 120 mg every 8 hours, diltiazem, 90 mg every 8 hours, and nifedipine, 20 mg every 8 hours, in randomized crossover fashion. Mean theophylline oral clearance decreased 18% and 12% after verapamil and diltiazem, respectively (p less than 0.05). No significant change in theophylline oral clearance was observed after nifedipine. Increases in mean theophylline half-life were observed after verapamil (10.8 +/- 3.2 hours) and diltiazem (9.9 +/- 2.4 hours) (p less than 0.05) but not after nifedipine (8.6 +/- 2.4 hours) when compared with control (8.6 +/- 1.9 hours). Apparent volume of distribution was unchanged. Urinary excretion data showed that the relative formation rate constants of 3-methylxanthine and 1,3-dimethyluric acid were decreased during verapamil and diltiazem (p less than 0.05) but not during nifedipine. No change in 1-methyluric acid excretion was observed. Increases in urinary elimination of unchanged theophylline were observed after verapamil, diltiazem, and nifedipine. The modest reduction in theophylline clearance observed after verapamil and diltiazem is not likely to produce clinically significant increases in theophylline concentrations in most patients.

Chloramphenicol sodium succinate kinetics in critically ill patients.

Chloramphenicol sodium succinate (SCAP) kinetics were studied in 10 critically ill patients. High-performance liquid chromatography was used to assay SCAP and chloramphenicol (CAP) in serum and urine. Total body (ClTB), metabolic (ClM), and renal (ClR) clearances of SCAP were variable. Correlations were found between creatinine clearance (Clcr) and ClTB, ClM, and ClR of SCAP (r = 0.92, p less than 0.001; r = 0.84, p less than 0.005; and r = 0.84, p less than 0.005). Recovery of SCAP in the urine also demonstrated large interpatient variability. Between 6.5% and 43.5% of the SCAP dose was recovered in the urine of 6 patients. This variability could not be explained by incomplete urine collection or by differences in renal function. Renal excretion of SCAP was shown to influence CAP serum levels. CAP ClTB was diminished, but no relationship was found between routine liver function studies and CAP ClTB. Therefore we caution the use of such relationships in using CAP in critically ill patients.

Effect of food on lidocaine kinetics: mechanism of food-related alteration in high intrinsic clearance drug elimination.

The effect of high-protein meal on the hepatic clearance (ClH) of intravenous lidocaine, because of its conceptual importance in understanding first-pass metabolic phenomena, was evaluated in nine healthy males. Our randomized crossover study demonstrated that mean ClH rose from 1245 to 1477 ml/min (P less than 0.03) as a result of the meal (i.e., mean area under the blood concentration-time curve decreased 20%). The magnitude of the change in clearance correlated weakly with fasting ClH (r = 0.54; slope = -0.037% per ml/min; intercept = 67.2%; P less than 0.15). In a separate study, it was observed that the meal did not influence lidocaine serum protein binding; the free fraction of lidocaine in samples drawn from the subjects in the fasting state averaged 0.305 +/- 0.027 while that from subjects who had eaten was 0.321 +/- 0.042. These data suggest that the mean clearance of lidocaine is increased by stimulation of hepatic blood flow rate. Furthermore, the magnitude of this increase is consistent with expectations based on a simple physiologic model. Thus, these data provide experimental support for the hypothesis that transient increases in splanchnic blood flow rate observed after a high-protein meal may explain apparent improvement of the oral bioavailability of model high intrinsic clearance drugs.

Effects of erythromycin or rifampin on losartan pharmacokinetics in healthy volunteers.

BACKGROUND: Losartan is metabolized by CYP2C9 and CYP3A4 to an active metabolite, E3174, which has greater antihypertensive activity than the parent compound. Coadministered drugs that inhibit or induce metabolic processes may therefore alter the pharmacokinetics and pharmacologic response of losartan and E3174. OBJECTIVE AND METHODS: Ten healthy volunteers were studied to assess the effects of CYP3A4 inhibition and nonspecific P450 enzyme induction on the pharmacokinetics of losartan and E3174. Subjects completed three 1-week phases separated by 6 days: 50 mg losartan every morning, losartan plus 500 mg erythromycin four times a day, and losartan plus 300 mg rifampin (INN, rifampicin) twice a day. On the eighth day of each phase, serial plasma concentrations of losartan and E3174 were obtained over 32 hours and steady-state pharmacokinetics were determined. RESULTS: Rifampin decreased the area under the concentration-time curve from time zero to 24 hours after the dose (AUC[0-24]) of losartan by 35% (349 +/- 246 versus 225 +/- 130; p = 0.0001) and decreased the AUC(0-24) of E3174 by 40% (1336 +/- 445 versus 792 +/- 302; p < 0.005). Losartan oral clearance was increased by 44% (p = 0.0001). The half-life values of both compounds were decreased by 50% (p < 0.005). In contrast, erythromycin did not significantly affect the AUC(0-24) or half-life of either losartan or E3174. CONCLUSIONS: Rifampin is a potent inducer of losartan and E3174 elimination. Given the magnitude of the effect, this interaction is likely to be clinically significant. On the basis of the minimal inhibitory effects observed with erythromycin, CYP3A4 appears to play a minor role in the in vivo metabolism of losartan to E3174. Further studies are needed to define the contribution of other isozymes, particularly CYP2C9, to the pharmacokinetics of losartan and E3174.

Altered protein binding of etoposide in patients with cancer.

Etoposide plasma protein binding (PB) is reported to be 94% based on in vitro studies using normal human serum albumin (SA). Etoposide PB in 17 patients with cancer receiving etoposide (50 to 100 mg/m2) and in plasma of 14 volunteers was determined by equilibrium dialysis with 3H-etoposide. The unbound fraction (Fu) in patients with cancer was 0.139 +/- 0.099 compared with 0.043 +/- 0.0036 in plasma from normal volunteers (p less than 0.0009; t test). Etoposide binding ratio (BR) was correlated directly with SA (r2 = 0.83; p less than 0.05). In the population with cancer Fu was significantly correlated with bilirubin (r2 = 0.837; p less than 0.05). In a multivariate analysis, SA and bilirubin were significant predictors of Fu (r2 = 0.93; p less than 0.05). This study corroborates previous reports of etoposide PB in normal human serum and demonstrates altered PB in patients with abnormal serum albumin or bilirubin levels.

Hepatic drug clearance in children with leukemia: changes in clearance of model substrates during remission-induction therapy.

We administered a "cocktail" of three model substrates for hepatic processes: antipyrine for oxidation, lorazepam for glucuronidation, and indocyanine green for hepatic blood flow, to children with acute lymphocytic leukemia (ALL). The plasma clearance of these substrates was determined before and after remission-induction therapy in 14 children with ALL. We found an improvement in clearance of antipyrine (0.65 to 0.95 ml/min/kg; P less than 0.01) and lorazepam (0.93 to 1.20 ml/min/kg; P less than 0.05) in 12 of 14 patients, with a mean increase of 67% and 52%, respectively, from before to after remission. There was no significant difference in mean indocyanine green clearance from before to after induction (14.8 vs. 14.4 ml/min/kg). There were no significant differences in liver function test results (SGOT, prothrombin time, or serum bilirubin) from before to after induction. Plasma concentrations of albumin, alpha 1-acid glycoprotein, and apolipoprotein A changed by a mean of +11.1%, -38.2%, and +68.6%, respectively, from before to after remission. However, these changes did not account for the changes in total plasma clearance of lorazepam, because lorazepam free fraction did not change and lorazepam free clearance increased by a mean of 83%. Our hypothesis is that eradication of hepatic leukemic infiltration by ALL remission therapy resulted in an improvement in microsomal metabolism of antipyrine and lorazepam.

Scaffolds for tissue engineering of cartilage.

Articular cartilage lesions resulting from trauma or degenerative diseases are commonly encountered clinical problems. It is well-established that adult articular cartilage has limited regenerative capacity, and, although numerous treatment protocols are currently employed clinically, few approaches exist that are capable of consistently restoring long-term function to damaged articular cartilage. Tissue engineering strategies that focus on the use of three-dimensional scaffolds for repairing articular cartilage lesions offer many advantages over current treatment strategies. Appropriate design of biodegradable scaffold conduits (either preformed or injectable) allow for the delivery of reparative cells bioactive factors, or gene factors to the defect site in an organized manner. This review seeks to highlight pertinent design considerations and limitations related to the development, material selection, and processing of scaffolds for articular cartilage tissue engineering, evidenced over the last decade. In particular, considerations for novel repair strategies that use scaffolds in combination with controlled release of bioactive factors or gene therapy are discussed, as are scaffold criteria related to mechanical stimulation of cell-seeded constructs. Furthermore, the subsequent impact of current and future aspects of these multidisciplinary scaffold-based approaches related to in vitro and in vivo cartilage tissue engineering are reported herein.

Heritability of nociception I: responses of 11 inbred mouse strains on 12 measures of nociception.

It is generally acknowledged that humans display highly variable sensitivity to pain, including variable responses to identical injuries or pathologies. The possible contribution of genetic factors has, however, been largely overlooked. An emerging rodent literature documents the importance of genotype in mediating basal nociceptive sensitivity, in establishing a predisposition to neuropathic pain following neural injury, and in determining sensitivity to pharmacological agents and endogenous antinociception. One clear finding from these studies is that the effect of genotype is at least partially specific to the nociceptive assay being considered. In this report we begin to systematically describe and characterize genetic variability of nociception in a mammalian species, Mus musculus. We tested 11 readily-available inbred mouse strains (129/J, A/J, AKR/J, BALB/cJ, C3H/HeJ, C57BL/6J, C58/J, CBA/J, DBA/2J, RIIIS/J and SM/J) using 12 common measures of nociception. These included assays for thermal nociception (hot plate, Hargreaves' test, tail withdrawal), mechanical nociception (von Frey filaments), chemical nociception (abdominal constriction, carrageenan, formalin), and neuropathic pain (autotomy, Chung model peripheral nerve injury). We demonstrate the existence of clear strain differences in each assay, with 1.2 to 54-fold ranges of sensitivity. All nociceptive assays display moderate-to-high heritability (h2 = 0.30-0.76) and mediation by a limited number of apparent genetic loci. Data comparing inbred strains have considerable utility as a tool for understanding the genetics of nociception, and a particular relevance to transgenic studies.

Heritability of nociception II. 'Types' of nociception revealed by genetic correlation analysis.

Clinical pain syndromes, and experimental assays of nociception, are differentially affected by manipulations such as drug administration and exposure to environmental stress. This suggests that there are different 'types' of pain. We exploited genetic differences among inbred strains of mice in an attempt to define these primary 'types'; that is, to identify the fundamental parameters of pain processing. Eleven randomly-chosen inbred mouse strains were tested for their basal sensitivity on 12 common measures of nociception. These measures provided for a range of different nociceptive dimensions including noxious stimulus modality, location, duration and etiology, among others. Since individual members of inbred strains are identical at all genetic loci, the observation of correlated strain means in any given pair of nociceptive assays is an index of genetic correlation between these assays, and hence an indication of common physiological mediation. Obtained correlation matrices were subjected to multivariate analyses to identify constellations of nociceptive assays with common genetic mediation. This analysis revealed three major clusters of nociception: (1) baseline thermal nociception, (2) spontaneously-emitted responses to chemical stimuli, and (3) baseline mechanical sensitivity and cutaneous hypersensitivity. Many other nociceptive parameters that might a priori have been considered closely related proved to be genetically divergent.