Inhibitory Effect Of Noradrenaline Research Articles

Spasmolytic Effect of Flopropione Does Not Involve Catechol-O- methyltransferase (COMT) Inhibition.

Flopropione (Flo) has been used for gallstone and urolithiasis as a spasmolytic agent almost exclusively in Japan. According to the package insert, its main mechanism is catechol-O-methyltransferase (COMT) inhibition and anti-serotonergic effect. This is obviously contrary to pharmacological common sense, but it is described that way in pharmacology textbooks and occurs in questions in the National Examination for Pharmacists in Japan. As this is a serious problem in education, we re-examined the action of Flo. The guinea pig ureter was hardly contracted by serotonin, but noradrenaline (NA) elicited repetitive twitch contraction, which was inhibited by Flo. The sphincter of Oddi (SO) exhibited a spontaneous repetitive twitch contraction, which was inhibited by NA and Flo. The inhibitory effect of NA was reversed by α- and β-blockers, whereas that of Flo was not. Entacapone, a representative COMT inhibitor, did not affect the movement of the ureter and the SO. Nifedipine suppressed carbachol-induced contraction of the taenia coli, spontaneous movement of the SO, and NA-induced contraction of the ureter to almost the same extent, whereas Flo did not inhibit the taenia coli, but inhibited the contraction of the SO and the ureter. The inhibitory pattern of Flo resembled that of the ryanodine receptor agonist 4-chloro-m-cresol and the inositol 1,4,5-trisphosphate (IP3) receptor antagonist 2-aminoethoxydiphenyl borate. It is concluded that COMT inhibition or serotonin inhibition is not involved in the spasmolytic action of Flo. Flo might act on ryanodine receptors and/or IP3 receptors, which are responsible for periodic Ca release from Ca stores, to disrupt coordinated Ca dynamics.

Activation of alpha-2 adrenergic receptors stimulates GABA release by astrocytes.

Norepinephrine is one of the key neurotransmitters in the hippocampus, but its role in the functioning of the neuroglial networks remains unclear. Here we show that norepinephrine suppresses NH4 Cl-induced oscillations of the intracellular Ca2+ concentration ([Ca2+ ]i ) in hippocampal neurons. We found that the inhibitory effect of norepinephrine against ammonium-induced [Ca2+ ]i oscillations is mediated by activation of alpha-2 adrenergic receptors. Furthermore, UK 14,304, an agonist of alpha-2 adrenergic receptors, evokes a biphasic [Ca2+ ]i elevation in a minor population of astrocytes. This elevation consists of an initial fast, peak-shaped [Ca2+ ]i rise, mediated by Giβγ subunit and subsequent PLC-induced mobilization of Ca2+ from internal stores, and a plateau phase, mediated by a Ca2+ influx from the extracellular medium through store-operated and TRPC3 channels. We show the correlation between the Ca2+ response in astrocytes and suppression of [Ca2+ ]i oscillations in neurons. The inhibitory effect of UK 14,304 is abolished in the presence of gallein, an inhibitor of Gβγ -signaling. In turn, application of the agonist in the presence of the PLC inhibitor decreases the frequency and amplitude of [Ca2+ ]i oscillations in neurons but does not suppress them. The same effect is observed in the presence of bicuculline, a GABA(A) receptor antagonist. We demonstrate that UK 14,304 application increases the frequency and amplitude of slow outward chloride currents in neurons, indicating the release of GABA by astrocytes. Thus, our findings indicate that the activation of astrocytic alpha-2 adrenergic receptors stimulates GABA release from astrocytes via Giβγ subunit-associated signaling pathway, contributing to the suppression of neuronal activity.

Presynaptic inhibition of transient receptor potential vanilloid type 1 (TRPV1) receptors by noradrenaline in nociceptive neurons.

The transient receptor potential vanilloid type 1 (TRPV1) receptor is a polymodal molecular integrator in the pain pathway expressed in Aδ- and C-fibre nociceptors and is responsible for the thermal hyperalgesia associated with inflammatory pain. Noradrenaline strongly inhibited the activity of TRPV1 channels in dorsal root ganglia neurons. The effect of noradrenaline was reproduced by clonidine and antagonized by yohimbine, consistent with contribution of α2 adrenergic receptors. The inhibitory effect of noradrenaline on TRPV1 channels was dependent on calcium influx and linked to calcium/calmodulin-dependent protein kinase II. In spinal cord slices, clonidine reduced the frequency of capsaicin-induced miniature EPSCs in the presence of tetrodotoxin and ω-conotoxin-MVIIC, consistent with inhibition of presynaptic TRPV1 channels by α2 adrenergic receptors. We suggest that modulation of presynaptic TRPV1 channels in nociceptive neurons by descending noradrenergic inputs may constitute a mechanism for noradrenaline to modulate incoming noxious stimuli in the dorsal horn of the spinal cord. The transient receptor potential vanilloid type 1 (TRPV1) receptor is a well-known contributor to nociceptor excitability. To address whether noradrenaline can down-regulate TRPV1 channel activity in nociceptors and reduce their synaptic transmission, the effects of noradrenaline and clonidine were tested on the capsaicin-activated current recorded from acutely dissociated small diameter (<27μm) dorsal root ganglia (DRG) neurons and on miniature (m)EPSCs recorded from large lamina I neurons in horizontal spinal cord slices. Noradrenaline or clonidine inhibited the capsaicin-activated current by ∼60%, and the effect was reversed by yohimbine, confirming that it was mediated by activation of α2 adrenergic receptors. Similarly, clonidine reduced the frequency of capsaicin-induced mEPSCs by ∼60%. Inhibition of capsaicin-activated current by noradrenaline was mediated by GTP binding proteins, and was highly dependent on calcium influx. The inhibitory effect of noradrenaline on the capsaicin-activated current was not affected either by blocking the activity of protein kinase A with H89, or by blocking the activity of protein kinase C with bisindolylmaleimide II. In contrast, when the calcium/calmodulin-dependent protein kinase II (CaMKII) was blocked with KN-93, the inhibitory effect of noradrenaline on the capsaicin-activated current was greatly reduced, suggesting that activation of adrenergic receptors in DRG neurons is preferentially linked to CaMKII activity. We suggest that modulation of TRPV1 channels by noradrenaline in nociceptive neurons is a mechanism whereby noradrenaline may suppress incoming noxious stimuli at the primary synaptic afferents in the dorsal horn of the spinal cord.

In vitro spontaneous contractile activity of colonic smooth muscle in naive Lewis rats: Acute effect of gonadal hormones.

Functional gastrointestinal disorders affect females more often. Changes in colonic motility may be etiological co-factors for the clinical symptoms. The aim of the present study was to analyze the influence of gonadal hormones on colonic contractile activity. In vitro measurements of colonic contractile activity in longitudinal smooth muscle strips of female and male Lewis rats were performed in an organ chamber experiment. After the administration of a gonadal hormone estradiol [EST], progesterone [PROG] and testosterone [TEST]) or ethanol solution as control, stimulation with acetylcholine (ACh) or inhibition with norepinephrine (NE) was performed. Compared to the smooth muscle strips of male rats, significantly higher spontaneous colonic contractile activity (SCCA) was observed in female animals. Increasing doses of ACh showed the progressive stimulation of SCCA whereas rising doses of NE resulted in a stepwise inhibition of SCCA, respectively. EST superfusion displayed an inhibitory effect on SCCA in both sexes and inhibited the ACh effect in female rats. Similarly, acute superfusion with high-dose PROG inhibited SCCA in females. Acute TEST superfusion inhibited SCCA in males and led to significant higher colonic contractile activity in males following subsequent stimulation with ACh. In female rats, the inhibitory effect of NE was reduced by prior exposure to TEST. In our in vitro study the acute exposure of colonic smooth muscle tissue to gonadal hormones led to sex-dependent changes in SCCA and translated in a modified response of smooth muscle strips to both pro-contractile and anti-contractile neurotransmitters.

Effect of Gonadal Hormones on Colonic Contractile Activity in Albino Rats

Sex differences were reported in Irritable Bowel Syndrome. We aimed to demonstrate the effect of gonadal hormones on colonic contractile activity. In this study, we performed in vitro measurements of colonic contractile activity in longitudinal smooth muscle strips of female and male Albino rats. After administration of a gonadal hormone (estrogen (EST), progesterone (PROG) and testosterone (TEST)) or ethanol (ETH) solution as control, stimulation with acetylcholine (ACh) or inhibition with norepinephrine (NE) was performed. We found that increasing doses of EST resulted in a stepwise inhibition of SCCA in both sexes and inhibited the ACh effect in females. Similarly, high dose PROG inhibited SCCA in females. TEST inhibited SCCA in males but not Ach induced contractions. TEST reduced the inhibitory effect of NE in female rats. In conclusion, exposure of colonic smooth muscle strips to gonadal hormones led to sex-dependent changes in SCCA and modified the response of smooth muscle strips to both pro-contractile and anti-contractile neurotransmitters.

Noradrenergic inhibition of spinal hyperexcitation elicited by cutaneous cold stimuli in rats with oxaliplatin-induced allodynia: electrophysiological and behavioral assessments.

We investigated the spinal action of noradrenaline on cold-elicited hyperexcitation detected in dorsal horn neurons of rats with allodynia induced by an oxaliplatin (6mg/kg, i.p.) injection. In vivo extracellular recordings from the spinal dorsal horn showed that wide dynamic range neurons responded to cutaneous acetone (10μl) stimulation in normal rats, and cold-elicited firings in oxaliplatin-administered rats were increased with a longer duration, correlated with behavioral responses. These responses were significantly attenuated by spinal administration (50μM) of noradrenaline or its agonists, clonidine (α2), phenylephrine (α1) and isoprenaline (β), in descending order of efficacy. Thus, the inhibitory effect of noradrenaline on spinal oxaliplatin-induced cold hyperexcitation is mediated mainly by activation of α2- and/or α1-adrenoceptors.

In vitro characterization of noradrenergic modulation of chemosensitive neurons in the retrotrapezoid nucleus.

Chemosensitive neurons in the retrotrapezoid nucleus (RTN) regulate breathing in response to CO2/H(+) changes and serve as an integration center for other autonomic centers, including brain stem noradrenergic neurons. Norepinephrine (NE) contributes to respiratory control and chemoreception, and, since disruption of NE signaling may contribute to several breathing disorders, we sought to characterize effects of NE on RTN chemoreception. All neurons included in this study responded similarly to CO2/H(+) but showed differential sensitivity to NE; we found that NE activated (79%), inhibited (7%), or had no effect on activity (14%) of RTN chemoreceptors. The excitatory effect of NE on RTN chemoreceptors was dose dependent, retained in the presence of neurotransmitter receptor blockers, and could be mimicked and blocked by pharmacological manipulation of α1-adrenergic receptors (ARs). In addition, NE-activation was blunted by XE991 (KCNQ channel blocker), and partially occluded the firing response to serotonin, suggesting involvement of KCNQ channels. However, in whole cell voltage clamp, activation of α1-ARs decreased outward current and conductance by what appears to be a mixed effect on multiple channels. The inhibitory effect of NE on RTN chemoreceptors was blunted by an α2-AR antagonist. A third group of RTN chemoreceptors was insensitive to NE. We also found that chemosensitive RTN astrocytes do not respond to NE with a change in voltage or by releasing ATP to enhance activity of chemosensitive neurons. These results indicate NE modulates subsets of RTN chemoreceptors by mechanisms involving α1- and α2-ARs.

Noradrenergic modulation of chemosensitive neurons in the retrotrapezoid nucleus

Chemosensitive neurons in the retrotrapezoid nucleus (RTN) regulate breathing in response to CO2/H+ changes, and serve as an integration center for other autonomic centers including brainstem noradrenergic neurons. Considering that application of norepinephrine (NE) into the RTN strongly modulates breathing (see abstract by Oliveira et al.) and disruption of NE signaling has been implicated in breathing disorders associated with Rett syndrome, sleep‐disordered breathing and multiple system atrophy, we sought to characterize the effects of NE on chemosensitive RTN neurons. Cell‐attached recordings were used to identify RTN chemoreceptors in brainstem slices (300 μm thick) isolated from rat pups (P7–11 days postnatal) by their characteristic response to CO2; i.e., low basal activity in 5% CO2 (0.2 ± 0.1 Hz) and high activity in 10% (1.8 ± 0.1 Hz) or 15% CO2 (2.1 ± 0.1 Hz ). We found that bath application of NE (1 μM) increased activity by 1.1 ± 0.1 Hz in 48 of 66 cells (73%), decreased activity by 1.0 ± 0.1 Hz in 6 of 66 cells (9%), or had no effect on activity in 12 of 66 (18%) of cells. Interestingly, basal activity but not CO2/H+‐sensitivity differed between each group based on their NE response; e.g., basal activity and firing response to 10% CO2 was 0.4 ± 0.1 and 1.2 ± 0.1 Hz (NE‐activated), 1.1 ± 0.3. and 1.1 ± 0.1 Hz (NE‐inhibited) and 0.03 ± 0.01 and 1.1 ± 0.01 Hz (NE‐insensitive). The excitatory effect of NE on RTN chemoreceptors was dose dependent (EC50 of 235 nM), retained during synaptic blockade (high Mg+2/low Ca2+) and could be mimicked by an α1‐receptor agonist (phenylephrine; 10 μM) and blocked with a α1‐receptor antagonist (prazosin; 1 μM). The inhibitory effect of NE on RTN chemoreceptors was retained during synaptic blockade and blunted by a α2‐receptor antagonist (idazoxan; 1 μM). A subset of NE‐activated cells were inhibited by NE in the presence of prazosin, whereas some NE‐inhibited cells showed an excitatory response to NE in the presence of idazoxan. A third group of RTN chemoreceptors did not respond to NE under control conditions or during hypercapnia, and these cells also did not respond to phenylephrine. In addition, whole‐cell voltage clamp (Vhold= −80 mV; TTX) recordings from CO2/H+‐sensitive RTN astrocytes show the absence of an NE‐sensitive current. These results indicate that i) RTN chemoreceptors are intrinsically sensitive to NE; ii) NE can increase or decrease chemoreceptor activity by α1‐ and α2‐receptor dependent mechanisms, respectively; and iii) NE modulates discrete subsets of RTN chemoreceptors based on basal activity and the differential contributions of α1‐α2‐receptors.Support or Funding InformationNIH Grant :HL104101

Anti-inflammatory effects of noradrenaline on LPS-treated microglial cells: Suppression of NFκB nuclear translocation and subsequent STAT1 phosphorylation

Noradrenaline (NA) has marked anti-inflammatory effects on activated microglial cells. The present study was conducted to elucidate the mechanisms underlying the NA effects using rat primary cultured microglial cells. NA, an α1 agonist, phenylephrine (Phe) and a β2 agonist, terbutaline (Ter) suppressed lipopolysaccharide (LPS)-induced nitric oxide (NO) release by microglia and prevented neuronal degeneration in LPS-treated neuron-microglia coculture. The agents suppressed expression of mRNA encoding proinflammatory mediators. Both an α1-selective blocker terazocine and a β2-selective blocker butoxamine overcame the suppressive effects of NA. cAMP-dependent kinase (PKA) inhibitors did not abolish the suppressive NA effects. LPS decreased IκB leading to NFκB translocation into nuclei, then induced phosphorylation of signal transducer and activator of transcription 1 (STAT1) and expression of interferon regulatory factor 1 (IRF1). NA inhibited LPS-induced these changes. When NFκB expression was knocked down with siRNA, LPS-induced STAT1 phosphorylation and IRF1 expression was abolished. NA did not suppress IL-6 induced STAT1 phosphorylation and IRF1 expression. These results suggest that one of the critical mechanisms underlying the anti-inflammatory effects of NA is the inhibition of NFκB translocation. Although inhibitory effects of NA on STAT1 phosphorylation and IRF1 expression may contribute to the overall suppressive effects of NA, these may be the downstream events of inhibitory effects on NFκB. Since NA, Phe and Ter exerted almost the same effects and PKA inhibitors did not show significant antagonistic effects, the suppression by NA might not be dependent on specific adrenergic receptors and cAMP-dependent signaling pathway.

Functional effects of β3-adrenoceptor on pacemaker activity in interstitial cells of Cajal from the mouse colon

We investigated the presence of β3-adrenoceptor and its functional effects on pacemaker potentials in colonic interstitial cells of Cajal (ICCs) from mice. The whole-cell patch clamp technique was used to record pacemaker potentials in cultured ICCs and reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the mRNA transcript levels β-adrenoceptors. The β3-adrenoceptor agonist, BRL37344, reduced the frequency of pacemaker potentials in a concentration-dependent manner. The inhibitory effects of BRL37344 were blocked by the pretreatment of propranolol, a nonspecific β-adrenoceptor antagonist, but not by the selective β1-adrenoceptor antagonist atenolol and the selective β2-adrenoceptor antagonist butoxamine. β3-adrenoceptor antagonists SR59230A and L748337 blocked the inhibitory effects of BRL37344. RT-PCR revealed mRNA transcripts of β1- and β3-adrenoceptor, but not β2-adrenoceptor, in c-kit- and Ano-1-positive colonic ICCs. The K+ channel blockers tetraethylammonium, apamin, and glibenclamide did not block the effects of BRL37344. Nω-Nitro-l-arginine methyl ester hydrochloride (L-NAME), an NO synthase inhibitor, and chelerythrine, a protein kinase C inhibitor, also did not block the effects of BRL37344. Noradrenaline mimicked the effects of BRL37344 in colonic ICCs. However, the inhibitory effects of noradrenaline on pacemaker potentials were blocked only by pretreatment with atenolol but not by butoxamine, SR59230A, or L748337. In small intestinal ICCs, BRL37344 had no effect on pacemaker potentials and mRNA transcripts of β1-and β2-adrenoceptor, but not β3-adrenoceptor were detected. These results suggest that β3-adrenoceptors are present in colonic ICCs and may play a role in regulating gastrointestinal motility by the inhibition of pacemaker potentials.

The functional activity of the adenylate cyclase system in the brains of rats with metabolic syndrome induced by immunization with peptide 11–25 of the type 4 melanocortin receptor

The melanocortin system of the brain, which includes melanocortin receptors of the fourth type (M4R), plays a key role in the regulation of energy homeostasis and controls functions of the nervous system. Inhibition of M4R results in obesity and the metabolic syndrome, which presumably occur due to changes in the neuromediator systems of the brain. To examine this hypothesis, we examined the effect of long-term immunization of rats with the BSA-conjugated K-[TSLHLWNRSSHGLHG11–25]-A peptide (K-[11–25]-A), which corresponds to the extracellular N-terminal domain of M4R, on the activity of the hormone-sensitive adenylate cyclase signaling system (ACSS) of the brain. In rats that were immunized numerous times with the BSA conjugate (the I group), we observed an increase in body weight, impaired glucose tolerance, insulin resistance, and dyslipidemia. At 13 months after the beginning of the experiment, we evaluated the ACSS activity in synaptosomal membranes from the brain. The basal activity of AC and its regulation by GppNHp and forskolin did not differ from the control. In the I group both the AC-stimulating effects of the α-melanocyte-stimulating hormone (α-MSH), M4R-agonist THIQ, dopamine, and pituitary AC-activating polypeptide and the AC-inhibiting effects of serotonin and 5-nonyloxytryptamine, an agonist of 5-hydroxytryptamine receptor (5-HTR) of the 1B/1D-subtype, decreased. The affinity of M4R to agonists did not change. The AC-stimulating effect of the M3R agonist γ-MSH was enhanced, which is a compensation for the weakening of M4R functions. The AC-stimulating effects of serotonin, EMD-386088, an agonist of 6 type 5-HTR relaxin, and noradrenaline, as well as the inhibitory effects of noradrenaline, the D2-agonist bromocriptine, and somatostatin in the I group did not change. Thus, inhibition of M4R as a result of immunization with the BSA conjugate of the K-[11–25]-A peptide alters the sensitivity of the ACSS of the rat brain to peptides of melanocortin family and other neurohormones. These alterations are characterized by hormonal and receptor specificity and may be one of the causes of insulin resistance, metabolic syndrome, and functional disturbances in the CNS and at the periphery under conditions of M4R deficit.

Abstract 253: Norepinephrine Suppresses T-lymphocyte Growth and Cytokine Production via Superoxide During Hypertension

Numerous studies have shown that pro-inflammatory cytokines produced by T-lymphocytes exacerbate hypertension. However, it is not clearly understood how endogenous pro-hypertensive stimuli such as norepinephrine (NE) directly act upon T-lymphocytes to modulate the responses of these inflammatory cells during hypertension. Herein, we hypothesized that increased systemic levels of NE directly activate T-lymphocytes during hypertension. To test this hypothesis, we examined the status of T-lymphocyte activation in the spleens of NE-infused hypertensive mice. Contrary to our hypothesis, while subcutaneous infusion of NE (3.8 μg/kg/min) significantly increased mean arterial pressure by approximately 18 ± 2 mmHg for 14-days (p&lt;0.05 vs. saline infused), splenic T-lymphocytes demonstrated no signs of increased activation. More specifically, splenic T-lymphocytes from NE-infused mice showed an approximate 20% ± 5% (p&lt;0.05) decrease in proliferation accompanied by a 50% ± 17% (p&lt;0.05) and 85% ± 6% (p&lt;0.05) reduction in interferon gamma (INFγ) and tumor necrosis factor alpha (TNFα) production respectively as compared to T-lymphocytes from saline-infused mice. Additionally, NE directly inhibited naïve T-lymphocyte proliferation and cytokine production ex vivo in a dose dependent manner. Furthermore, while NE did not demonstrate any pro-apoptotic effects on T-lymphocytes, a 21% ± 2% (p&lt;0.05 vs. saline) increase in G1 arrest was observed in NE-treated T-lymphocytes, and this was accompanied by a 60% ± 4% (p&lt;0.05 vs. saline) decrease in pro-growth cyclin D3, E1, and E2 mRNA expression. Interestingly, NE caused a 71% ± 17% (p&lt;0.05 vs. saline) increase in cellular superoxide (O 2 •- ) production as evidenced by dihydroethidium (DHE) oxidation, which was shown to be partially-causal to the inhibitory effects of NE as the addition of Tempol, a O 2 •- scavenger, to the drinking water of NE-infused mice was able to moderately restore T-lymphocyte growth and pro-inflammatory cytokine production. Taken together with previous studies, our data indicates that direct NE stimulation of naïve T-lymphocytes inhibits their proliferation and cytokine production in hypertension.

Noradrenaline stimulates cell proliferation by suppressing potassium channels via Gi/o‐protein‐coupled α1B‐adrenoceptors in human osteoblasts

Recent studies demonstrated that the sympathetic nervous system regulates bone metabolism via β(2) -adrenoceptors. Although α-adrenoceptors are also expressed in osteogenic cells, their functions in bone metabolism have been less studied. We previously demonstrated that noradrenaline suppressed potassium currents via α(1B) -adrenoceptors in the human osteoblast SaM-1 cell line. The aim of this study was to investigate the signal transduction pathway and the physiological role of noradrenaline in human osteoblasts in more detail. To investigate signal transduction through α(1B) -adrenoceptors, we used whole-cell patch clamp recording and Ca fluorescence imaging. Potassium channels regulate membrane potential and cell proliferation activity in non-excitable cells, so we evaluated cell proliferation activity by BrdU incorporation and WST assay. In SaM-1 cells, bath-applied noradrenaline elevated intracellular Ca(2+) concentration and this effect was abolished by both chloroethylclonidine, an α(1B) -adrenoceptor antagonist, and U73122, a PLC inhibitor. However, the inhibitory effect of noradrenaline on whole-cell current was unaffected by U73122. In contrast, in cells pretreated with either Pertussis toxin, a G(i/o) -protein-coupled receptor inhibitor, or gallein, a Gβγ-protein inhibitor, the inhibitory effect of noradrenaline on whole-cell current was significantly suppressed. Noradrenaline-induced enhancement of cell proliferation was inhibited by CsCl, a non-selective potassium channel blocker, gallein and H89, a PKA inhibitor, but not by U73122. Noradrenaline facilitated cell proliferation by regulation of potassium currents in human osteoblasts via G(i/o) -protein-coupled α(1B) -adrenoceptors, not via coupling to Gq-proteins.

Influence of exercise on NA- and Hsp72-induced release of IFNγ by the peritoneal suspension of macrophages and lymphocytes from genetically obese Zucker rats

Regular physical exercise is recognized as a nonpharmacological therapeutic strategy in the treatment of metabolic syndrome, and has been proposed for improving obesity, diabetic status, insulin resistance, and immune response. The aim of the present study was to evaluate the effect of a regular exercise program (treadmill running, 5days/week for 14weeks at 35cm/s for 35min in the last month) on the release of the pro-inflammatory cytokine interferon gamma (IFNγ) by peritoneal cells (macrophages and lymphocytes) from obese Zucker rats (fa/fa) in response to noradrenaline (NA) and heat shock proteins of 72kDa (Hsp72), and the possible adaptation due to training for a bout acute exercise (a single session of 25-35min at 35cm/s). In healthy (lean Fa/fa) and obese animals, peritoneal cells released greater concentrations of IFNγ in response to Hsp72 and lower concentrations in response to NA. The regular exercise training protocol, evaluated in the obese animals, produced a clear change in the regulation of the release of IFNγ. Peritoneal immune cells from trained animals released more IFNγ in response to NA, but there was a reduction in the release of IFNγ in response to Hsp72. In the obese animals, regular exercise caused a change in the inhibitory effect of NA (which now becomes stimulatory) and the stimulatory effect of Hsp72e (which now becomes inhibitory) in relation to the release of IFNγ. This reflects that Hsp72, induced by the prior release of NA following exercise-induced stress, plays a role in the homeostatic balance of release of IFNγ by peritoneal immune cells in obese animals during exercise.

Norepinephrine reduces ω-conotoxin-sensitive Ca2+ currents in renal afferent neurons in rats

Sympathetic efferent and peptidergic afferent renal nerves likely influence hypertensive and inflammatory kidney disease. Our recent investigation with confocal microscopy revealed that in the kidney sympathetic nerve endings are colocalized with afferent nerve fibers (Ditting T, Tiegs G, Rodionova K, Reeh PW, Neuhuber W, Freisinger W, Veelken R. Am J Physiol Renal Physiol 297: F1427-F1434, 2009; Veelken R, Vogel EM, Hilgers K, Amman K, Hartner A, Sass G, Neuhuber W, Tiegs G. J Am Soc Nephrol 19: 1371-1378, 2008). However, it is not known whether renal afferent nerves are influenced by sympathetic nerve activity. We tested the hypothesis that norepinephrine (NE) influences voltage-gated Ca(2+) channel currents in cultured renal dorsal root ganglion (DRG) neurons, i.e., the first-order neuron of the renal afferent pathway. DRG neurons (T11-L2) retrogradely labeled from the kidney and subsequently cultured, were investigated by whole-cell patch clamp. Voltage-gated calcium channels (VGCC) were investigated by voltage ramps (-100 to +80 mV, 300 ms, every 20 s). NE and appropriate adrenergic receptor antagonists were administered by microperfusion. NE (20 μM) reduced VGCC-mediated currents by 10.4 ± 3.0% (P < 0.01). This reduction was abolished by the α-adrenoreceptor inhibitor phentolamine and the α(2)-adrenoceptor antagonist yohimbine. The β-adrenoreceptor antagonist propranolol and the α(1)-adrenoceptor antagonist prazosin had no effect. The inhibitory effect of NE was abolished when N-type currents were blocked by ω-conotoxin GVIA, but was unaffected by other specific Ca(2+) channel inhibitors (ω-agatoxin IVA; nimodipine). Confocal microscopy revealed sympathetic innervation of DRGs and confirmed colocalization of afferent and efferent fibers within in the kidney. Hence NE released from intrarenal sympathetic nerve endings, or sympathetic fibers within the DRGs, or even circulating catecholamines, may influence the activity of peptidergic afferent nerve fibers through N-type Ca(2+) channels via an α(2)-adrenoceptor-dependent mechanism. However, the exact site and the functional role of this interaction remains to be elucidated.

Pre- and Post-junctional α-Adrenoceptor-mediated Responses in the Rat Gastric Fundus In-vitro

The effects of alpha 1- and alpha 2-adrenoceptor agonists on smooth muscle tone and on cholinergic excitatory and non-adrenergic, non-cholinergic inhibitory responses to field stimulation have been investigated in the rat gastric fundus in-vitro. None of the alpha-adrenoceptor agonists tested, noradrenaline, phenylephrine, cirazoline, guanoxabenz or UK-14,304 showed any contractile effects at concentrations up to 30 microM. In preparations where tone was raised by barium (0.5-2 mM), the mixed alpha 1- and alpha 2-adrenoceptor agonist noradrenaline (0.01-10 microM), and the selective alpha 1-adrenoceptor agonists cirazoline (0.01-10 microM) and phenylephrine (0.01-10 microM) produced concentration-dependent relaxations which were antagonized by the alpha 1-adrenoceptor antagonist prazosin (0.01-1.0 microM). The selective alpha 2-adrenoceptor agonists UK-14,304 (0.03-30 microM) and guanoxabenz (0.03-30 microM), had no relaxant effects in raised tone. UK-14,304 (0.03-1.0 microM) produced a concentration-dependent inhibition of cholinergic nerve-induced responses which was antagonized by the alpha 2-adrenoceptor antagonist idazoxan (0.03-1.0 microM) but not by prazosin (0.03-1.0 microM). Noradrenaline (0.03-1.0 microM) also produced an inhibition of cholinergic nerve-induced responses which was antagonized by idazoxan (0.03-1.0 microM). A small component of the noradrenaline inhibitory effects was antagonized by prazosin (10%). Cirazoline (0.03-1.0 microM) produced a small inhibition of cholinergic nerve-induced responses which was antagonized by prazosin (0.03-1.0 microM). The prazosin-sensitive components of the inhibitory effects of noradrenaline and cirazoline occurred at concentrations which also produced post-junctional relaxation.(ABSTRACT TRUNCATED AT 250 WORDS)

Different sites of action for alpha 2-adrenoceptor antagonists in the modulation of noradrenaline release and contraction response in the vas deferens of the rat.

Rat vas deferens was prepared, loaded with [3H]noradrenaline, and superfused to measure the release of tritium in resting conditions and in response to electrical field stimulation. The alpha 2-adrenoceptor antagonists yohimbine, CH-38083 (7,8-(methylenedioxi)-14 alpha-hydroxyalloberbane HCl), and idazoxan increased the electrically induced release of tritium in a concentration-dependent manner, whereas noradrenaline and the alpha 2-adrenoceptor agonist xylazine exerted opposite effects. The inhibitory effect of noradrenaline on electrically induced tritium release was antagonized by yohimbine, CH-38083, and idazoxan. Of the alpha 2-adrenoceptor antagonists tested, yohimbine and CH-38083 reversed the xylazine-induced inhibition of tritium release, and idazoxan was found to be completely ineffective against xylazine. Idazoxan, yohimbine and CH-38083 antagonized the inhibitory effect of xylazine on electrical stimulation-induced contractions of the vas deferens, as was evidenced by the apparent pA2 values. We conclude from the present experiments that noradrenaline and xylazine inhibit noradrenaline release by acting on distinct prejunctional alpha 2-adrenoceptors and that the receptor subtype that responds to xylazine is insensitive to idazoxan. In addition, inhibition by xylazine of contractility but not of noradrenaline release was antagonized by idazoxan, suggesting that besides noradrenergic neurotransmission, other motor transmitter systems (purinergic) may also be involved in the inhibition by alpha 2-adrenoceptor antagonists of mechanical responses in the rat vas deferens.

Inhibitory effects of sympathomimetic drugs on cholinergically mediated contractions of guinea-pig isolated tracheal muscle.

The experiments examine the actions of sympathomimetic drugs on the responses evoked by electrical field stimulation or by acetylcholine in guinea-pig tracheal strip chains. Electrical field stimulation evoked contractions which were cholinergically mediated, in the presence of guanethidine (10 microM) and indomethacin (2 microM). All the sympathomimetic drugs tested caused a concentration-dependent reduction in the height of these contractions. Inhibitory effects of isoprenaline and terbutaline were largely prevented by propranolol (2 microM) alone, whereas those of clonidine, oxymetazoline, lidamidine and WHR1370 were prevented by yohimbine alone (2 microM). Treatments with both propranolol and yohimbine were required to prevent the inhibitory effects of noradrenaline, adrenaline and dopamine. Contractions evoked by exogenous acetylcholine (0.1-3 microM) were also inhibited by all catecholamines and terbutaline, but not by clonidine, oxymetazoline, lidamidine and WHR1370. The inhibitory effects were antagonized by propranolol (2 microM) alone. The results suggest that in guinea-pig isolated tracheal muscle, sympathomimetic drugs can inhibit cholinergic neurotransmission not only by postjunctional beta 2-adrenoceptors but also by prejunctional alpha 2-adrenoceptors.

The effects of norepinephrine on lactoperoxidase enzyme (LPO)

Norepinephrine, a hormone and a neurotransmitter, was known as catecholamine. In the present study, we examined the inhibitory effect of norepinephrine on lactoperoxidase enzyme purified from bovine milk. Lactoperoxidase (LPO; E.C.1.11.1.7) was purified from bovine milk with three purification steps: Amberlite CG-50 resin, CM-Sephadex C-50 ion-exchange chromatography and Sephadex G-100 gel filtration chromatography, respectively. LPO was purified with a yield of 31.5%, a specific activity of 30.33 EU/mg proteins and 20.77 purification fold. To determine enzyme purity, SDS-PAGE was performed and single band was observed. The Rz (A412/A280) value for LPO was 0.9. The effect of norepinephrine on lactoperoxidase was determined using ABTS as a chromogenic substrate. The half maximal inhibitory concentration (IC50) value norepinephrine was found to be 67.2 µM. Also, inhibition constant (Ki) for norepinephrine was found to be 62.0 µM. Norepinephrine was found as non-competitive inhibitor. Key words: Noradrenaline, norepinephrine, lactoperoxidase, LPO, enzyme purification, inhibition.

Noradrenergic neurons expressing Fos during waking and paradoxical sleep deprivation in the rat

Noradrenaline is known to induce waking (W) and to inhibit paradoxical sleep (PS or REM). Both roles have been exclusively attributed to the noradrenergic neurons of the locus coeruleus (LC, A6), shown to be active during W and inactive during PS. However, the A1, A2, A5 and A7 noradrenergic neurons could also be responsible. Therefore, to determine the contribution of each of the noradrenergic groups in W and in PS inhibition, rats were maintained in continuous W for 3 h in a novel environment or specifically deprived of PS for 3 days, with some of them allowed to recover from this deprivation. A double immunohistochemical labeling with Fos and tyrosine hydroxylase was then performed. Thirty percent of the LC noradrenergic cells were found to be Fos-positive after exposure to the novel environment and less than 2% after PS deprivation. In contrast, a significant number of double-labeled neurons (up to 40% of the noradrenergic neurons) were observed in the A1/C1, A2 and A5 groups, after both novel environment and PS deprivation. After PS recovery and in control condition, less than 1% of the noradrenergic neurons were Fos-immunoreactive, regardless of the noradrenergic group. These results indicate that the brainstem noradrenergic cell groups are activated during W and silent during PS. They further suggest that the inhibitory effect of noradrenaline on PS may be due to the A1/C1, A2 and to a lesser degree to A5 neurons but not from those of the LC as previously hypothesized.