The selective MMP-12 inhibitor, AS111793 reduces airway inflammation in mice exposed to cigarette smoke.

BACKGROUND: Macrophage elastase (MMP-12) is involved in the inflammatory process of chronic obstructive pulmonary disease (COPD). The aim of this study was to investigate in mice the effect of MMP-12 inhibition on the inflammatory process induced by cigarette smoke (CS) or by lipopolysaccharide (LPS) exposure of the airways. EXPERIMENTAL APPROACH: C57BL/6 mice were given, orally, either the selective MMP-12 inhibitor AS111793 (3, 10, 30 and 100 mg kg(-1)), the PDE-4 inhibitor roflumilast (3 mg kg(-1)) or vehicle, then exposed to CS (for 3 days) or to LPS (100 microg mL(-1), 30 min). Subsequent to the last smoke or LPS exposure, bronchoalveolar lavages (BAL) were performed and lungs were removed and homogenized to analyze various markers of inflammation at appropriate times. KEY RESULTS: Inhibition of MMP-12 by AS111793 (10 and 30 mg kg(-1)) was associated with a reduction of the increase in neutrophil number in BAL fluids after 4 days and of macrophages after 11 days. On day 4, AS111793 also significantly reduced all the inflammation markers that had increased after CS exposure, including soluble TNF receptors I and II, MIP-1gamma, IL-6 and pro-MMP-9 activity in BAL fluids, and KC/CXCL1, fractalkine/CX3CL1, TIMP-1 and I-TAC/CXCL11 in lung parenchyma. In contrast, inhibition of MMP-12 did not reduce neutrophil influx, pro-MMP-9 activity or KC/CXCL1 release in BAL fluids of mice exposed to LPS. CONCLUSION: Inhibition of MMP-12 with AS111793, reduced the inflammatory process associated with exposure of mice to CS, strongly suggesting a specific involvement of MMP-12 in lung inflammation following CS exposure.

AS601245, a c-Jun NH2-terminal kinase (JNK) inhibitor, reduces axon/dendrite damage and cognitive deficits after global cerebral ischaemia in gerbils.

BACKGROUND AND PURPOSE: Based on their proven ability, in animal models of stroke, to reduce damage to brain grey matter, many drugs have been tested in clinical trials but without success. Failure to save axons from injury and to protect functional outcome has been proposed as the major reason for this lack of success. We have previously demonstrated in two rodent models of cerebral ischaemia, that AS601245 (1,3-benzothiazol-2-yl (2-([2-(3-pyridinyl) ethyl] amino)-4 pyrimidinyl) acetonitrile), an inhibitor of the c-Jun NH(2)-terminal kinase (JNK), has neuroprotective properties. The aim of the present study was to further investigate if AS601245 in addition to its ability to protect neurons also could protect neurites and preserve memory after cerebral ischaemia, in gerbils. EXPERIMENTAL APPROACH: Using immunohistochemical techniques and a behavioural test, we studied the effect of the compound AS601245 on neurodegeneration and cognitive deficits after global cerebral ischaemia in gerbils. KEY RESULTS: At a dose of 80 mg kg(-1), i.p., AS601245 reduced damage to neurites by 67% (P<0.001 versus controls) and activation of astrocytes by 84% (P<0.001 versus controls). In addition, AS601245 (80 mg kg(-1), i.p.) prevented ischaemia-induced impairment of memory in the inhibitory avoidance task model. CONCLUSIONS AND IMPLICATIONS: The present results suggest that AS601245 reduced damage to neurites and decreased astrogliosis following global ischaemia and also improved long-term memory, supporting JNK inhibition as a promising therapeutic strategy for ischaemic insults to the CNS.

Decrease in the reactivity of locus coeruleus neurons to hypotension after an increase in their tyrosine hydroxylase content: a subregional in vivo voltammetry study in the rat.

The aim of the present work was to determine if noradrenergic neurons of the anterior and the posterior subregions of the locus coeruleus exhibit a difference in reactivity in response to sodium nitroprusside-induced arterial hypotension, and if the pharmacological induction of tyrosine hydroxylase by RU24722 modifies the reactivity of locus coeruleus neurons to this hypotensive stimulus. Previous findings have demonstrated that administration of RU24722 increases the concentration of tyrosine hydroxylase in the rat locus coeruleus by two different mechanisms in the anterior and in the posterior locus coeruleus subregions. The goal of the present study was to measure in vivo the changes in catecholaminergic metabolism in the locus coeruleus after treatment with RU24722 using differential normal pulse voltammetry (DNPV). In vehicle-treated rats, arterial hypotension increased catecholaminergic metabolism with the same pattern in the two locus coeruleus subregions. However, the changes in the magnitude of the catechol oxidation current throughout the recording period were significantly smaller in the posterior subregion (P < 0.001). In the RU24722-pretreated rats, there was a 39% increase in tyrosine hydroxylase and dihydroxyphenylacetic acid in the locus coeruleus. The functional reactivity to hypotension measured by DNPV was significantly decreased (P < 0.001) in both the anterior and posterior locus coeruleus subregions with RU24722 treatment. Therefore, this study suggests that the response of locus coeruleus cells to a hypotensive stimulus depends upon the intracellular tyrosine hydroxylase concentration both in the basal condition and during pharmacological induction of tyrosine hydroxylase gene expression.

Changes in catecholamine metabolism in the rostral ventrolateral medulla following halothane and nitroprusside-induced hypotension: an in vivo electrochemical study.

The objective was to observe changes in rostral ventrolateral medulla (RVLM) catecholamine metabolism using in vivo voltammetry following induced hypotension with halothane or nitroprusside (SNP). Rats anesthetized (halothane, metocurine) and ventilated were stereotaxically implanted with carbon microelectrodes in the RVLM. The catechol oxidation current (CA.OC, % baseline) was used as an index of RVLM catecholaminergic metabolism. Groups of rats (n = 5) were given (A) halothane 0.75% for 60 min; (B) halothane 2.75% plus phenylephrine infusion to maintain mean arterial pressure (MAP) for 30 min, then halothane 0.75% for 30 min; (C) halothane (2.5-3.0%) for 30 min (MAP 60 +/- 5 mmHg) then halothane 0.75% for 30 min; (D) halothane 0.75% and sodium nitroprusside (SNP) for 30 min (MAP 60 +/- 5 mmHg), then halothane 0.75% for 30 min. Halothane 0.75% produced no significant change in CA.OC or MAP (A), while halothane 2.5-3.0% produced a significant decrease in MAP and a symmetrical significant increase in CA.OC (ANOVA, P less than 0.5). This increase peaked at 30 min (180 +/- 28%) and reached 110 +/- 9% baseline at 60 min. The halothane and phenylephrine combination produced no significant change in CA.OC or MAP during the 30 min exposure (B). SNP (D) produced a significant increase in CA.OC (peak 48 min, 224 +/- 35%) which remained elevated at 60 min (198 +/- 32%). Thus, the induced hypotension produced activation of RVLM catecholaminergic neurons. SNP induced a prolonged significant increase in RVLM catecholamine metabolism which may relate to rebound hypertension following use of this drug.

In vivo monitoring of catecholaminergic metabolism in the C1 region of rat medulla oblongata: a comparative study by voltammetry and intracerebral microdialysis.

In vivo voltammetry or microdialysis was used to monitor catecholaminergic metabolism in the C1 region of the ventrolateral medulla oblongata of anesthetized rats. In vivo voltammetry allowed the recording of a catechol oxidation current (CA.OC) peak in this region. This CA.OC was suppressed after inhibition of monoamine oxidase by pargyline or after inhibition of tyrosine hydroxylase by alpha-methyl-p-tyrosine and was markedly increased after blockade of dopamine-beta-hydroxylase by FLA 63. Similar results were found when intracerebral microdialysis coupled with HPLC and electrochemical detection was used to measure the concentration of 3,4-dihydroxyphenylacetic acid (DOPAC) in the dialysates obtained from the C1 region: The changes in CA.OC and DOPAC concentration in the dialysates exhibited very similar kinetic characteristics in the three pharmacological experiments. These results support the involvement of DOPAC as a major component of the electrochemical signal recorded by voltammetry in the C1 group of adrenergic neurons.

Pharmacological and functional evidence for extracellular 3,4-dihydroxyphenylacetic acid as an index of metabolic activity of the adrenergic neurons: an in vivo voltammetry study in the rat rostral ventrolateral medulla.

Catecholamine metabolism was studied in vivo in the C1 adrenergic area of the rostral ventrolateral medulla oblongata in rats, using differential normal pulse voltammetry coupled with an activated carbon fiber microelectrode. Pharmacological evidence indicates that 3,4-dihydroxyphenylacetic acid, the major dopamine metabolite, is responsible for the electrochemical signal appearance in the C1 group, and that it reflects the catecholamine synthesis rate, as previously reported in the locus coeruleus. Indeed, 3,4-dihydroxyphenylacetic acid was estimated to be formed from 77% of the intracellular dopamine, since its synthesis was increased by only 23%, after blockade of the dopamine-beta-hydroxylase activity. Neuronal activation by retrograde electrical stimulation increased the electrochemical signal, as well as hemorrhage and hypotension, suggesting that the level of extracellular 3,4-dihydroxyphenylacetic acid is a good biochemical index of the C1 adrenergic cellular activity in baseline conditions and during cellular activation.

Continuous volume infusion improves circulatory stability in anesthesized rats.

'Optimized' management (OM) was provided to chloralose-anesthetized rats for 12 h by combining continuous infusion (7 ml.kg-1). mechanical ventilation and strict control of acid-base equilibrium (n = 7). The chloralose-anesthetized rats managed conventionally (conventional management: CM, n = 9) received neither volume infusion, nor mechanical ventilation, nor correction of acid-base disturbances. All the OM rats completed the study while 6 out of 9 CM rats died before the end of the study period. Mean arterial pressure (MAP) remained at 100 mmHg for 12 h in the OM group. MAP stayed close to 70 mmHg in the CM group for 6 h and declined to very low levels thereafter (mean +/- S.E.M.: 46.0 +/- 3.9 mmHg at 12 h, P less than 10(-4) when compared to the other group). Central venous pressure and cardiac output remained close to baseline values for 12 h in the OM group. Acid-base equilibrium was preserved in the OM group in contrast to a severe metabolic acidosis in the CM group (pH = 7.14 +/- 0.03 at 12 h; P less than 10(-4). Such as 'optimized' management involving mechanical ventilation with oxygen, continuous infusion and acid-base monitoring may be of value to maintain circulatory stability in anesthetized rodent preparations during long periods of time, as in neurophysiological experiments.

Fentanyl decreases catecholamine metabolism measured by in vivo voltammetry in the rat locus coeruleus.

The objective of this study was to investigate under controlled conditions the effects of fentanyl on the rat locus coeruleus catechol oxidation current. Using differential normal pulse voltammetry combined with electrochemically treated carbon fiber electrodes to measure the catechol oxidation current, catecholamine metabolism can be reliably monitored. Male Sprague-Dawley rats weighing 500-600 g had carbon fiber electrodes implanted into the locus coeruleus under halothane - O2 - air anesthesia with controlled ventilation and muscle relaxation. Experiments consisted of four groups of rats given the following treatments: (A) saline (n = 6); (B) fentanyl, 10 micrograms.kg-1 i.v. (n = 6); (C) naloxone, 800 micrograms.kg-1 i.v. followed 2 min later by fentanyl, 10 micrograms.kg-1 (n = 5); (D) clonidine, 200 micrograms.kg-1 i.p. (n = 6). There was no significant change in the catechol oxidation current following saline. Fentanyl produced a significant (ANOVA, p less than 0.05) decrease in the catechol oxidation current (maximum 32 min postinjection was 75.8 +/- 4.6% of baseline). This decrease was prevented by a prior injection of naloxone. Clonidine produced a significant decrease in catechol oxidation current (maximum 40 min postinjection was 54.1 +/- 7.0% of baseline). Systolic blood pressure was significantly decreased following clonidine and there were no significant changes in arterial blood gases throughout the experiments. The alpha 2-adrenergic agonist clonidine and the opioid fentanyl produced a decrease in locus coeruleus catechol oxidation current measured by in vivo voltammetry, which monitors catecholamine turnover.

Subcellular distribution of tyrosine hydroxylase in some catecholaminergic rat brain areas determined by a quantitative immunoblot assay.

The subcellular distribution of the protein tyrosine hydroxylase (TH) after fractionation of rat brain tissue was studied by a sensitive technique of immunoblot quantification in the dopaminergic nigrostriatal and the dorsal noradrenergic pathways and in the ventrolateral medulla. This repartition indicates that in all catecholaminergic regions of the cell bodies studied, the contribution of the nerve endings to the total TH amount is very low (less than 7%), in contrast to that observed in the terminal fields. The correlative subcellular determination of the TH amount and activity in the same tissue could be a useful approach for studying experimentally induced mechanisms of catecholamine synthesis modulation in different brain catecholaminergic pathways.

Baroreflex-linked variations of catecholamine metabolism in the caudal ventrolateral medulla: an in vivo electrochemical study.

In vivo electrochemical recordings of the metabolism of catecholamines were obtained in the caudal ventrolateral medulla in anesthetized rats submitted to various experimental changes in systemic arterial pressure. Hypertension induced with phenylephrine and reversal of hypovolemia decreased the catechol metabolic activity. In contrast, controlled or hypovolemic hypotension, induced respectively with sodium nitroprusside or blood withdrawal (30% of blood volume), reversibly elicited the opposite pattern. This was suppressed by deafferentation. The changes in catechol metabolic activity in response to hypovolemia were accompanied by similar trends of variations of plasma vasopressin levels. By contrast with the increased catechol metabolic activity secondary to hypotension induced by either prazosin, sodium nitroprusside or hypovolemia, clonidine elicited a decrease in catechol metabolic activity. These data show a dynamic and specific involvement of the metabolism of catecholamines themselves promoted by changes in systemic arterial pressure. This pattern of functioning of catechol metabolism in the caudal ventrolateral medulla appears to be negatively related to systemic arterial pressure changes, a finding which does not fit with the proposed vasodepressor role of the A1-group.

Clonidine modulates the ventrolateral medullary catechol metabolic hyperactivity induced by hypotension.

In vivo electrochemistry allowed recording of a catechol oxidation current in the ventrolateral medulla, caudal to the obex, in anesthetized rats whose ventilatory, metabolic and hemodynamic parameters were rigorously controlled. Hemorrhage or controlled hypotension induced an increase in the metabolism of catecholamines in the A1 noradrenergic group, which remained activated after full hemodynamic recovery. Clonidine (200 micrograms.kg-1 i.p.) given 30 min prior to hemorrhage or immediately before controlled hypotension suppressed partially the increased metabolism of catecholamines especially during the recovery period. This suggests that clonidine preserved phasic reactivity upon circulatory disturbances and decreased tonic hyperactivity following circulatory recovery.