Teleantagonism: A pharmacodynamic property of the primary nociceptive neuron

Previous work from our group showed that intrathecal (i.t.) administration of substances such as glutamate, NMDA, or PGE(2) induced sensitization of the primary nociceptive neuron (PNN hypernociception) that was inhibited by a distal intraplantar (i.pl.) injection of either morphine or dipyrone. This pharmacodynamic phenomenon is referred to in the present work as ""teleantagonism``. We previously observed that the antinociceptive effect of i.t. morphine could be blocked by injecting inhibitors of the NO signaling pathway in the paw (i.pl.), and this effect was used to explain the mechanism of opioid-induced peripheral analgesia by i.t. administration. The objective of the present investigation was to determine whether this teleantagonism phenomenon was specific to this biochemical pathway (NO) or was a general property of the PNNs. Teleantagonism was investigated by administering test substances to the two ends of the PNN (i.e., to distal and proximal terminals; i.pl. plus i.t. or i.t. plus i.pl. injections). We found teleantagonism when: (i) inhibitors of the NO signaling pathway were injected distally during the antinociception induced by opioid agonists; (ii) a nonselective COX inhibitor was tested against PNN sensitization by IL-1 beta; (iii) selective opioid-receptor antagonists tested against antinociception induced by corresponding selective agonists. Although the dorsal root ganglion seems to be an important site for drug interactions, the teleantagonism phenomenon suggests that, in PNNs, a local sensitization spreads to the entire cell and constitutes an intriguing and not yet completely understood pharmacodynamic property of this group of neurons.

Role of complement C5a in mechanical inflammatory hypernociceptio: potential use of C5a receptor antagonists to control inflammatory pain

particularly neutrophil chemoattraction. Herein, the role of C5a in the genesis of inflammatory hypernociception was investigated in rats and mice using the specific C5a receptor antagonist PMX53 (AcF-[OP(D-Cha)WR]). Experimental approach: Mechanical hypernociception was evaluated with a modification of the Randall-Selitto test in rats and electronic pressure meter paw test in mice. Cytokines were measured by ELISA and neutrophil migration was determined by myeloperoxidase activity. Key results: Local pretreatment of rats with PMX53 (60-180 mg per paw) inhibited zymosan-, carrageenan-, lipopolysaccharide (LPS)- and antigen-induced hypernociception. These effects were associated with C5a receptor blockade since PMX53 also inhibited the hypernociception induced by zymosan- activated serum and C5a but not by the direct-acting hypernociceptive mediators, prostaglandin E-2 and dopamine. Underlying the C5a hypernociceptive mechanisms, PMX53 did not alter the cytokine release induced by inflammatory stimuli. However, PMX53 inhibited cytokine-induced hypernociception. PMX53 also inhibited the recruitment of neutrophils induced by zymosan but not by carrageenan or LPS, indicating an involvement of neutrophils in the hypernociceptive effect of C5a. Furthermore, the C5a-induced hypernociception was reduced in neutrophil-depleted rats. Extending these findings in rats, blocking C5a receptors also reduced zymosan- induced joint hypernociception in mice. Conclusions and implications: These results suggest that C5a is an important inflammatory hypernociceptive mediator, acting by a mechanism independent of hypernociceptive cytokine release, but dependent on the presence of neutrophils. Therefore, we suggest that inhibiting the action of C5a has therapeutic potential in the control of inflammatory pain.

Dorsal raphe nucleus regulation of a panic-like defensive behavior evoked by chemical stimulation of the rat dorsal periaqueductal gray matter

Electrical or chemical stimulation of the dorsal periaqueductal gray matter (DPAG) evokes escape, a defensive behavior that has been related to panic attacks. Injection of 5-HT(1A) or 5-HT(2A) receptor agonists into this midbrain area inhibits this response. It has been proposed that the impairment of 5-HT mechanisms controlling escape at the level of the DPAG may underlie the susceptibility to panic attacks that characterizes the panic disorder. In this study we evaluated the effects of the pharmacological manipulation of the dorsal raphe nucleus (DRN), which are the main source of 5-HT input to the DPAG, on the escape response evoked in rats by the intra-DPAG injection of the nitric oxide donor SIN-1. The results showed that DRN administration of the 5-HT(1A) receptor agonist 8-OH-DPAT which inhibits the activity of 5-HT neurons favored the expression of escape induced by SIN-1. Intra-DRN injection of the excitatory amino acid kainic acid or the 5-HT(1A) receptor antagonist WAY-100635 did not change escape expression. However, both compounds fully blocked the escape reaction generated by intra-DPAG injection of the excitatory amino acid D,L-homocysteic acid (DLH). Overall, the results indicate that 5-HT neurons in the DRN exert a bidirectional control upon escape behavior generated by the DPAG. Taking into account the effect of WAY-100635 on DLH-induced escape, they also strengthen the view that DRN 5-HT(1A) autoreceptors are under tonic inhibitory influence by 5-HT. (C) 2010 Elsevier B.V. All rights reserved.

IL-17 mediates articular hypernociception in antigen-induced arthritis in mice

IL-17 is an important cytokine in the physiopathology of rheumatoid arthritis (RA). However, its participation in the genesis of nociception during RA remains undetermined. In this study, we evaluated the role of IL-17 in the genesis of articular nociception in a model of antigen (mBSA)-induced arthritis. We found that mBSA challenge in the femur-tibial joint of immunized mice induced a dose-and time-dependent mechanical hypernociception. The local IL-17 concentration within the mBSA-injected joints increased significantly over time. Moreover, co-treatment of mBSA challenged mice with an antibody against IL-17 inhibited hypernociception and neutrophil recruitment. In agreement, intraarticular injection of IL-17 induced hypernociception and neutrophil migration, which were reduced by the pre-treatment with fucoidin, a leukocyte adhesion inhibitor. The hypernociceptive effect of IL-17 was also reduced in TNFR1(-/-) mice and by pre-treatment with infliximab (anti-TNF antibody), a CXCR1/2 antagonist or by an IL-1 receptor antagonist. Consistent with these findings, we found that IL-17 injection into joints increased the production of TNF-alpha, IL-1 beta and CXCL1/KC. Treatment with doxycycline (non-specific MMPs inhibitor), bosentan (ET(A)/ET(B) antagonist), indomethacin (COX inhibitor) or guanethidine (sympathetic blocker) inhibited IL-17-induced hypernociception. IL-17 injection also increased PGE(2) production, MMP-9 activity and COX-2, MMP-9 and PPET-1 mRNA expression in synovial membrane. These results suggest that IL-17 is a novel pro-nociceptive cytokine in mBSA-induced arthritis, whose effect depends on both neutrophil migration and various pro-inflammatory mediators, as TNF-alpha, IL-1 beta, CXCR1/2 chemokines ligands, MMPs, endothelins, prostaglandins and sympathetic amines. Therefore, it is reasonable to propose IL-17 targeting therapies to control this important RA symptom. (C) 2009 International Association for the Study of Pain. Published by Elsevier B. V. All rights reserved.

Acute and persistent nociceptive paw sensitisation in mice: The involvement of distinct signalling pathways

Aims: Many fundamental pharmacological studies in pain and inflammation have been performed on rats. However, the pharmacological findings were generally not extended to other species in order to increase their predictive therapeutic value. We studied acute and chronic inflammatory nociceptive sensitisation of mouse hind paws by prostaglandin E(2) (PGE(2)) or dopamine (DA), as previously described in rats. We also investigated the participation of the signalling pathways in acute and persistent sensitisation. Main methods: Mechanical sensitisation (hypernociception) induced by intraplantar administrations of PGE(2) or DA was evaluated with an electronic pressure meter. The signalling pathways were pharmacologically investigated with the pre-administration of adenylyl cyclase (AC), cAMP-dependent protein kinase (PKA), protein kinase C epsilon (PKC epsilon), and the extracellular signal-related kinase (ERK) inhibitors. Key findings: Single or 14 days of successive intraplantar injections of PGE(2) or DA-induced acute and persistent hypernociception (lasting for more than 30 days), respectively. The involvement of AC, PKA or PKC epsilon was observed in the acute hypernociception induced by PGE(2), while PKA or PKC epsilon were continuously activated during the period of persistent hypernociception. The acute hypernociception induced by DA involves activation of ERK, PKC epsilon, AC or PKA, while persistent hypernociception implicated ERK activation, but not PKA, PKC epsilon or AC. Significance: In mice, acute and persistent paw sensitisation involves the different activation of kinases, as previously described for rats. This study opens the possibility of comparing pharmacological approaches in both species to further understand acute and chronic inflammatory sensitisation, and possibly associated genetic manipulations. (C) 2009 Elsevier Inc. All rights reserved.

Effect of baroreceptor denervation on the autonomic control of arterial pressure in conscious mice

This study evaluated the role of arterial baroreceptors in arterial pressure (AP) and pulse interval (PI) regulation in conscious C57BL mice. Male animals, implanted with catheters in a femoral artery and a jugular vein, were submitted to sino-aortic (SAD), aortic (Ao-X) or carotid sinus denervation (Ca-X), 5 daysprior to the experiments. After basal recording of AP, the lack of reflex bradycardia elicited by administration of phenylephrine was used to confirm the efficacy of SAD, and cardiac autonomic blockade with methylatropine and propranolol was performed. The AP and PI variability were calculated in the time and frequency domains (spectral analysis/fast Fourier transform) with the spectra quantified in low-(LF; 0.25-1Hz) and high-frequency bands (HF; 1-5Hz). Basal AP and AP variability were higher after SAD, Ao-X or Ca-X than in intact mice. Pulse interval was similar among the groups, whereas PI variability was lower after SAD. Atropine elicited a slight tachycardia in control mice but did not change PI after total or partial denervation. The bradycardia caused by propranolol was higher after SAD, Ao-X or Ca-X compared with intact mice. The increase in the variability of AP was accompanied by a marked increase in the LF and HF power of the AP spectra after baroreceptor denervation. The LF and HF power of the PI were reduced by SAD and by Ao-X or Ca-X. Therefore, both sino-aortic and partial baroreceptor denervation in mice elicits hypertension and a remarkable increase in AP variability and cardiac sympathetic tonus. Spectral analysis showed an important contribution of the baroreflex in the power of LF oscillations of the PI spectra. Both sets of baroreceptors seem to be equally important in the autonomic regulation of the cardiovascular system in mice.

Role of the spinal cord NO/cGMP pathway in the control of arterial pressure and heart rate

The modulatory effect of nitric oxide/cyclic guanosine monophosphate (NO/cGMP) pathway on sympathetic preganglionic neurons still deserves further investigation. The present study was designed to examine the role of the spinal cord NO/cGMP pathway in controlling mean arterial pressure and heart rate. We observed that intrathecal administration of the NO synthase inhibitor N omega-Nitro-L-arginine methyl ester hydrochloride (L-NAME) causes an increase in mean arterial pressure but does not affect heart rate. Intrathecal administration of the soluble guanylyl cyclase inhibitor 1H-[1,2,4] Oxadiazolo[4,3-a] quinoxalin-1-one (ODQ) does not change mean arterial pressure and heart rate. The precursor for NO synthesis, L-arginine, reduces both mean arterial pressure and heart rate while administration of ODQ before L-arginine impaired decreases in mean arterial pressure and heart rate. Administration of the N-methyl-D-aspartate (NMDA) receptor antagonist DL-2-amino-5-phosphonopentanoic acid (AP5) after L-NAME does not affect increases in mean arterial pressure promoted by NO synthase inhibition. Although the hypotensive and bradycardic responses induced by intrathecal administration of L-arginine depend on cGMP, our results indicate that NO acts to tonically inhibit SPNs, independent of either cGMP or NMDA receptors.

GABA and nitric oxide in the PVN are involved in arterial pressure control but not in the chemoreflex responses in rats

GABAergic, nitrergic and glutamatergic mechanisms in the PVN on the baseline mean arterial pressure (MAP), heart rate (HR) and on the cardiovascular responses to chemoreflex activation in awake rat were evaluated. Chemoreflex was activated with KCN before and after microinjections into the PVN. Bicuculline into the PVN increased baseline MAP (94+/-3 vs 113+/-5 mmHg) and HR (350+/-9 vs 439+/-18 bpm) but had no effect on the pressor (49+/-5 vs 47+/-6 mmHg) or bradicardic (-213+/-23 vs -256+/-42 bpm) responses (n=7). Kynurenic acid into the PVN (n=6) produced no significant changes in the MAP (98+/-3 vs 100+/-3 mmHg), HR (330+/-5 vs 339+/-12 mmHg) or in the pressor (50+/-4 vs 42+/-4 mmHg) and bradicardic (-252+/-4 vs -285+/-16 bpm) responses to chemoreflex. L-NAME into the PVN (n=8) produced increase in the MAP (94+/-3 vs 113+/-5 mmHg) and HR (350+/-9 vs 439+/-18 bpm) but had no effect on the pressor (52+/-5 vs 47+/-6 mmHg) or bradicardic (-253+/-19 vs -320+/-25 bpm) responses to chemoreflex. We conclude that GABA(A) and nitric oxide in the PVN are involved in the maintenance of the baseline MAP but not in the modulation of the responses to chemoreflex. The results also show that Glutamate receptors in the PVN are not involved in maintenance of the baseline MAP, HR or in the cardiovascular responses to chemoreflex in awake rats. (C) 2008 Elsevier B.V. All rights reserved.

Changes in hemodynamic and neurohumoral control cause cardiac damage in one-kidney, one-clip hypertensive mice

Borges GR, Salgado HC, Silva CA, Rossi MA, Prado CM, Fazan R Jr. Changes in hemodynamic and neurohumoral control cause cardiac damage in one-kidney, one-clip hypertensive mice. Am J Physiol Regul Integr Comp Physiol 295: R1904-R1913, 2008. First published October 1, 2008; doi:10.1152/ajpregu.00107.2008.-Sympathovagal balance and baroreflex control of heart rate (HR) were evaluated during the development (1 and 4 wk) of one-kidney, one-clip (1K1C) hypertension in conscious mice. The development of cardiac hypertrophy and fibrosis was also examined. Overall variability of systolic arterial pressure (AP) and HR in the time domain and baroreflex sensitivity were calculated from basal recordings. Methyl atropine and propranolol allowed the evaluation of the sympathovagal balance to the heart and the intrinsic HR. Staining of renal ANG II in the kidney and plasma renin activity (PRA) were also evaluated. One and four weeks after clipping, the mice were hypertensive and tachycardic, and they exhibited elevated sympathetic and reduced vagal tone. The intrinsic HR was elevated only 1 wk after clipping. Systolic AP variability was elevated, while HR variability and baroreflex sensitivity were reduced 1 and 4 wk after clipping. Renal ANG II staining and PRA were elevated only 1 wk after clipping. Concentric cardiac hypertrophy was observed at 1 and 4 wk, while cardiac fibrosis was observed only at 4 wk after clipping. In conclusion, these data further support previous findings in the literature and provide new features of neurohumoral changes during the development of 1K1C hypertension in mice. In addition, the 1K1C hypertensive model in mice can be an important tool for studies evaluating the role of specific genes relating to dependent and nondependent ANG II hypertension in transgenic mice.

Infection control practices among a cohort of Brazilian dentists

Objective: To evaluate infection control practices among dentists in private and public practice. Design: Survey and cross-sectional analysis. Setting: Sertaozinho city, Brazil. Participants: All dentists who were currently working at the study city, and agreed to participate, resulting in a study population of 135 dentists. Methods: Participants were personally interviewed and variables were submitted to X(2) or Fisher`s exact test. Results: Hand washing before and after each patient was reported by 86.7% of dentists, but private practitioners used liquid soap and paper towels more often than their public colleagues (p<0.001). Most of the study population (97.8%) used gloves routinely during clinical sessions, but 8.2% reused them. Dry-heat was the main method employed for sterilisation of heat-stable devices by 80.0% of dentists, but adequate temperature and time of exposure was accomplished by only 32.1% of public and 70.0% of private professionals (p<0.001). Heat-sensitive devices were disinfected with an adequate substance by 60.0% of both affiliation dentists (p=0.908). Conclusions: There is a large gap between infection control recommendations and practices observed among the study population, and the situation is worse in public services. To reverse that situation, infection control issues must be openly debated by professional associations, dental schools and health authorities.

The effect of aerobic physical training on cardiac autonomic control of rats submitted to ovariectomy

Objective: To investigate the effect of aerobic physical training on cardiovascular autonomic control in ovariectomized rats using different approaches. Design: Female Wistar rats were divided into four groups: sedentary sham rats (group SSR), trained sham rats (group TSR), sedentary ovariectomized rats (group SOR), and trained ovariectomized rats (group TOR). Animals from the trained groups were submitted to a physical training protocol (swimming) for 12 weeks. Results: Pharmacological evaluation showed that animals from group TSR had an increase in their cardiac vagal tonus compared with the animals from groups SSR and SOR. The analysis of heart rate variability (HRV) showed that groups TSR and SOR had fewer low-frequency oscillations (0.20-0.75 Hz) compared with groups SSR and TOR. When groups TSR and SOR were compared, the former was found to have fewer oscillations. With regard to high-frequency oscillations (0.75-2.5 Hz), group SSR had a reduction compared with the other groups, whereas group TSR had the greatest oscillation compared with groups SOR and TOR, with all values expressed in normalized units. Analysis of HRV was performed after pharmacological blockade, and low-frequency oscillations were found to be predominantly sympathetic in sedentary animals, whereas there was no predominance in trained animals. Conclusion: Ovariectomy did not change the tonic autonomic control of the heart and, in addition, reduced the participation of sympathetic component in cardiac modulation. Physical training, on the other hand, increased the participation of parasympathetic modulation on the HRV, including ovariectomized rats.

The effect of ovariectomy on cardiac autonomic control in rats submitted to aerobic physical training

We have investigated the ovariectomy effects on the cardiovascular autonomic adaptations induced by aerobic physical training and the role played by nitric oxide (NO). Female Wistar rats (n =70) were divided into five groups: Sedentary Sham (SS): Trained Sham (TS); Trained Hypertensive Sham treated with N(C)-nitro-L-arginine methyl ester (L-NAME) (THS): Trained Ovariectomized (TO); and Trained Hypertensive Ovariectomized treated with L-NAME (THO). Trained groups were submitted to a physical training during 10 weeks. The cardiovascular autonomic control was investigated in all groups using different approaches: 1) pharmacological evaluation of autonomic tonus with methylatropine and propranolol; 2) analysis of heart rate (HR) and systolic arterial pressure (AP) variability; 3) spontaneous baroreflex sensitivity (BRS) evaluation. Hypertension was observed in THS and THO groups. Pharmacological analysis showed that TS group had increased predominance of autonomic vagal tonus compared to SS group. HR and intrinsic HR were found to be reduced in all trained animals. TS group, compared to other groups, showed a reduction in LF oscillations (LF=0.2-0.75 Hz) of pulse interval in both absolute and normalized units as well as an increase in HF oscillations (HF=0.75-2.50 Hz) in normalized unit. FIRS analysis showed that alpha-index was different between all groups. TS group presented the greatest value, followed by the TO, SS. THO and THS groups. Ovariectomy has negative effects on cardiac autonomic modulation in trained rats, which is characterized by an increase in the sympathetic autonomic modulation. These negative effects suggest NO deficiency. In contrast, the ovariectomy seems to have no effect on AP variability. (C) 2008 Elsevier B.V. All rights reserved.

Inflammatory mediators involved in the nociceptive and oedematogenic responses induced by Tityus serrulatus scorpion venom injected into rat paws.

The present study investigated the role of kinins, prostaglandins (PGs) and nitric oxide (NO) in mechanical hypernociception, Spontaneous nociception and paw oedema after intraplantar (ipl) injection of Tityus serrulatus venom (Tsv) in male Wistar rats. Tsv was ipl-injected in doses of 0.01-10 mu g/paw. Pre-treatment (30 min prior) with DALBK (100 nmol/paw) and icatibant (10 nmol/paw), B1 and B2 selective kinin receptor antagonists, L-NAME (50 mg/kg, i.p., a non-selective nitric oxide synthase inhibitor) or celecoxib, selective COX-2 inhibitor, was given 1 h prior per os (5 mg/kg, p.o.), significantly reduced the hypernociceptive response (Von Frey method), the spontaneous nociception (determined by counting the number of flinches) and paw oedema (plethysmometer method) induced by Tsv at doses of 1.0 and 10 mu g/paw for both nociceptive and oedematogenic responses, respectively. Nevertheless, indomethacin (5 mg/kg, i.p.. 30 min prior) was ineffective in altering all of these events. The results of the present study show that Tsv, injected ipl into the rat paw, causes a dose-dependent paw oedema, mechanical hypernociception and flinches (a characteristic biphasic response) in which kinins and NO are substantially involved. Although celecoxib was effective against the oedema and pain caused by Tsv, COX-2 does not seem to be involved in the inflammatory response caused by Tsv. (C) 2008 Elsevier Ltd. All rights reserved.

Fibrinolytic activity is associated with presence of cystic medial degeneration in aneurysms of the ascending aorta

Fibrinolytic activity is associated with presence of cystic medial degeneration in aneurysms of the ascending aorta Aims: Thoracic ascending aortic aneurysms (TAA) are characterized by elastic fibre breakdown and cystic medial degeneration within the aortic media, associated with progressive smooth muscle cell (SMC) rarefaction. The transforming growth factor (TGF)-beta/Smad2 signalling pathway is involved in this process. Because the pericellular fibrinolytic system activation is able to degrade adhesive proteins, activate matrix metalloproteinase (MMP), induce SMC disappearance and increase the bioavailability of TGF-beta, the aim was to investigate the plasminergic system in TAA. Methods and results: Ascending aortas [21 controls and 19 TAAs (of three different aetiologies)] were analysed. Immunohistochemistry showed accumulation of t-PA, u-PA and plasmin in TAAs, associated with residual SMCs. Overexpression of t-PA and u-PA was confirmed by reverse transcription-polymerase chain reaction (RT-PCR), immunoblotting and zymography on TAA extracts and culture medium conditioned by TAA. Plasminogen was present on the SMC surface and inside cytoplasmic vesicles, but plasminogen mRNA was undetectable in the TAA medial layer. Plasmin-antiplasmin complexes were detected in TAA-conditioned medium and activation of the fibrinolytic system was associated with increased fibronectin turnover. Fibronectin-related material was detected immunohistochamically in dense clumps around SMCs and colocalized with latent TGF-beta binding protein-1. Conclusions: The fibrinolytic pathway could play a critical role in TAA progression, via direct or indirect impact on ECM and consecutive modulation of TGF-beta bioavailability.

Multiplex semi-nested RT-PCR with exogenous internal control for simultaneous detection of bovine coronavirus and group A rotavirus

Neonatal calf diarrhea is a multi-etiology syndrome of cattle and direct detection of the two major agents of the syndrome, group A rotavirus and Bovine coronavirus (BCoV) is hampered by their fastidious growth in cell culture. This study aimed at developing a multiplex semi-nested RT-PCR for simultaneous detection of BCoV (N gene) and group A rotavirus (VP1 gene) with the addition of an internal control (mRNA ND5). The assay was tested in 75 bovine feces samples tested previously for rotavirus using PAGE and for BCoV using nested RT-PCR targeted to RdRp gene. Agreement with reference tests was optimal for BCoV (kappa = 0.833) and substantial for rotavirus detection (kappa = 0.648). the internal control, ND5 mRNA, was detected successfully in all reactions. Results demonstrated that this multiplex semi-nested RT-PCR was effective in the detection of BCoV and rotavirus, with high sensitivity and specificity for simultaneous detection of both viruses at a lower cost, providing an important tool for studies on the etiology of diarrhea in cattle. (C) 2010 Elsevier B.V. All rights reserved.