Inability of an opioid antagonist lacking negative intrinsic activity to induce opioid receptor up-regulation in vivo.

1. It has recently been suggested that opioid antagonists may be divided into those possessing negative intrinsic activity (e.g. naloxone) and those with neutral intrinsic activity (e.g. MR2266). 2. MR2266 was chronically administered to rats by subcutaneous infusion at a dose of 0.3 mg kg-1 h-1 for 1 week. 3. This dose reduced ingestive behaviour and blocked the antinociceptive effects of a kappa-agonist, indicating occupation of opioid receptors in vivo. 4. No supersensitivity could be detected to the antinociceptive actions of mu or kappa agonists, either one or two days after cessation of treatment. 5. No up-regulation of mu, delta or kappa binding sites was observed. 6. Since naloxone induces both supersensitivity and receptor up-regulation under equivalent conditions, the results suggest that negative intrinsic activity may be required for these phenomena to occur.     

Centrally administered opioid antagonists, nor-binaltorphimine, 16-methyl cyprenorphine and MR2266, suppress intake of a sweet solution

Three opioid antagonists (MR2266, 16-methyl cyprenorphine and nor-binaltorphimine) were tested independently for their ability to suppress the intake of a highly palatable saccharin and glucose (S/G) solution after central administration. MR2266 is an equally potent antagonist at kappa (kappa) and mu (mu) opioid receptors. Nor-binaltorphimine (N-BNI) and 16-methyl cyprenorphine (M80) are two recently developed opioid antagonists that were chosen based upon their ability to act more selectively than naloxone at kappa and delta (delta) opioid receptor types, respectively. Prior research has demonstrated that when dissolved in acid and administered centrally, MR2266 (20 micrograms) fails to suppress S/G intake. Because all three antagonists are rather insoluble in water, they were dissolved in dimethyl sulfoxide (DMSO). Rats with chronic ventricular cannula were allowed to consume S/G for a 0.5 hr bout. They received a single intracerebroventricular (ICV) injection of antagonist (MR2266: 0, 10, 20 and 40 micrograms; M80: 0, 5, 10, 20 and 40 micrograms or N-BNI: 0, 1, 3, and 10 micrograms) 10 min prior to the start of the drinking bout. Administration of DMSO alone failed to alter drinking relative to saline, whereas each antagonist significantly attenuated S/G intake. We conclude that, when dissolved in DMSO, these antagonists suppress drinking by blockade of opioid receptors.     

An analysis of precipitated withdrawal in rats acutely dependent on morphine

Acute dependence on a single dose of morphine was assessed in Wistar rats by observing the frequencies of occurrence of several signs of withdrawal precipitated by naloxone, diprenorphine, Mr2097, Mr1452 and Mr2266. Naloxone significantly precipitated urination, paw shakes, head shakes and chewing. Diprenorphine significantly precipitated urination and chewing. Mr2097 precipitated urination, head shakes, teeth chattering and chewing. The selective kappa antagonists Mr1452 and Mr2266 significantly precipitated only urination and teeth chattering. Signs of the precipitated withdrawal by Mr2097 were mediated by stereoselective opioid receptors, as the other diastereoisomer, Mr2097, did not precipitate them. Stereospecific opioid receptors were also involved in the induction of acute dependence, as naloxone precipitated withdrawal only in I-methadone-treated rats, but not in d-methadone treated rats. All the opioid antagonists produced at least some degree of "abstinoid" signs in morphine-free rats which might be caused by the blockade of endogenous opioids acting on mu and/or kappa receptors. The signs of withdrawal precipitated by naloxone and Mr2097 might be primarily mediated by mu receptors, those of diprenorphine by both mu and kappa receptors, and those by Mr1452 and Mr2266 were likely to be selectively mediated by kappa receptors. The latter aspect was further supported by experiments showing that the novel kappa agonist U-50488H did not precipitate withdrawal. A low degree of precipitation of withdrawal by Mr1452 and Mr2266 and the absence of precipitation of abstinence by U-50488H might be related to either a lack or an existence of a low proportion of kappa receptors in rat brain. Further experiments using selective agonists and antagonists are needed to evaluate these findings.     

Tifluadom-induced diuresis in rats. Evidence for an opioid receptor-mediated central action

Tifluadom dose-dependently induced diuresis in rats after subcutaneous injection and oral application. (+)Tifluadom was at least 100-fold more potent than the (-)enantiomer in inducing diuresis. The diuretic action of tifluadom was dose-relatedly reduced by the opioid receptor antagonists naloxone and MR 2266. Naloxone methobromide did not antagonize the diuretic effect of tifluadom nor did the benzodiazepine receptor blocker Ro 15-1788. These data demonstrate that the diuretic effect of tifluadom is mediated centrally via an agonistic interaction between the drug and opioid receptors.     

Characterization of kappa opioid receptors in the rabbit ear artery

Quantitative characterization of the kappa opioid receptor in the rabbit ear artery was carried out using three kappa-selective agonist compounds, dynorphin-(1-13), U-69593 and ethylketocyclazocine. Kinetic analysis was performed using the antagonist, MR 2266. Two other in vitro preparations were studied for comparison: the mouse was deferens and rabbit was deferens. To avoid mu receptor action in the mouse was deferens the irreversible mu receptor antagonist, beta-funaltrexamine, was used. It was demonstrated that, using the highly selective kappa agonist compound U-69593, Ke values for MR 2266 obtained in the three assay systems were not significantly different. These results suggest that kappa receptors present in these three tissues share identical properties.     

Vagally mediated reflex and cardiac slowing induced by loperamide in rats

The intravenous injection of loperamide induced an immediate fall in blood pressure and heart rate in anaesthetized rats. Both effects were inhibited by the opiate antagonists naloxone and MRZ 2266 BS. Bilateral vagotomy also inhibited both effects whereas atropine only reduced the bradycardia, but the combination of atropine and tertatolol suppressed the bradycardia. A high dose of loperamide induced bradycardia in pithed rats. This effect was prevented by MRZ 2266 BS but not by naloxone. It is concluded that loperamide can elicit a vagally mediated reflex involving vagal and sympathetic mechanisms and could stimulate cardiac opiate receptors, probably kappa, both effects leading to bradycardia.     

Ethylketocyclazocine and bremazocine analgesia in neonatal rats

In three experiments we examined the analgesic potency of kappa opioid receptor agonists in 2- and 16-day-old rats. Ethylketocyclazocine (1–50 mg/kg) produced similar dose- and time-dependent increases in the latency to retract a hind paw from a noxious thermal stimulus in rats of both ages. Bremazocine (0.001–10 mg/kg), a kappa agonist with reported antagonist activity at mu receptors, was also effective in producing analgesia in 2-day-old rats. The dose-effect relationship for bremazocine was nonmonotonic. Bremazocine analgesia (0.1 mg/kg) was reversed by both naltrexone and MR2266, a putative kappa opioid antagonist. These results are discussed in terms of the functional integrity of a kappa analgesic system in the developing rat.     

On the mechanism of anticonvulsant effect of tramadol in mice

The present study was conducted to examine the effects of tramadol, an atypical opioid on convulsive behaviour in maximal electroshock (MES) seizure test on mice. Moreover, an attempt was also made to investigate the role of possible receptor mechanisms involved. MES seizures were induced via transauricular electrodes (60 mA, 0.2 sec). Seizure severity was determined by (1) the duration of tonic hindlimb extensor (THE) phase and by (2) mortality due to electroconvulsions. Intraperitoneal (i.p.) administration of tramadol dose-dependently (10-50 mg/kg) decreased the duration of THE phase of MES. The anticonvulsant effect of tramadol was antagonized by the opioid antagonists, naloxone in high dose, and MR2266, a selective kappa antagonist but not by naltrindole, a delta opioid antagonist. Coadministration of either gamma-aminobutyric acid (GABA)-ergic drugs (diazepam, GABA, muscimol and baclofen) or N-methyl-D-aspartate (NMDA) receptor antagonist, MK801 with tramadol augmented the anticonvulsant effect of the latter drug. By contrast, flumazenil, a central benzodiazepine (BZD) receptor antagonist, counteracted the diazepam-induced facilitation of anti-MES effect of tramadol. Similarly, delta-aminovaleric acid (DAVA), a GABAB receptor antagonist, abolished the facilitatory effect of baclofen, a GABAB agonist on anti-MES action of tramadol. These BZD-GABAergic antagonists, flumazenil or DAVA, on their own also antagonized the anti-MES effect of tramadol administered alone. No significant effect on mortality was observed in any of the studied groups. Taken together, the current results have demonstrated a possible role for multitude of important neurotransmitter systems, i.e., opioid (kappa), GABAA-BZD receptors system, GABAB receptors and NMDA channel involvement in the antielectroshock effect of tramadol in mice.     

Effect of opioid antagonists on acetylcholine release from guinea-pig brain slices: influence of peptidase inhibition

With the aim of ascertaining the existence of an endogenous opioid control on cholinergic structures, the effects of the opioid antagonists naloxone, Mr 1452, Mr 2266, and ICI 174864 on spontaneous and electrically evoked [3H]choline (Ch) efflux from guinea-pig brain slices were tested. In cerebral cortex and caudate nucleus slices, no drug changed either resting or stimulus-evoked 3H-Ch efflux whether in the absence or in the presence of peptidase inhibitors (thiorphan 0.3 microM, bestatin 10 microM, captopril 10 microM, l-leucyl-l-leucine 2 mM). Conversely, in thalamus slices, the benzomorphan opioid antagonists Mr 1452 and Mr 2266 (but not their non-opioid stereoisomers Mr 1453 and Mr 2267) dose dependently increased St2/St1 ratios, when applied 15 min before St2 to slices superfused with normal Krebs solution. Peptidase inhibition potentiated the facilitatory effect of Mr compounds. Peptidase inhibitors per se reduced the stimulus evoked efflux of 3H-Ch, when present either from the beginning or from 15 min before St2 and potentiated the inhibition induced by dynorphin (Dyn). Mr 2266 fully antagonized any inhibitory effect induced by peptidase inhibitors, Dyn or their combination. ICI 174864, a selective delta antagonist, did not affect 3H-Ch efflux, while naloxone showed a tendency to increase it when peptidases were inhibited. Taken together, these data suggest the existence of an endogenous opioid tone inhibiting acetylcholine release in the guinea-pig thalamus. The relative affinities of the antagonists used suggest that the receptor involved may be of the kappa type.     

Studies on the anticonvulsant effect of U50488H on maximal electroshock seizure in mice

The present study was designed to investigate the effect of U50488H, a prototype non-peptide kappa opioid agonist on convulsive behaviour using a maximal electroshock (MES) seizure test in mice. An attempt was also made to explore the role of possible receptors involved. MES seizures were induced via transauricular electrodes (60 mA, 0.2 s). Seizure severity was evaluated by means of two parameters, i.e., (1). duration of tonic hindlimb extensor phase and (2). mortality due to convulsions. Intraperitoneal (i.p.) administration of U50488H dose dependently (5-20 mg/kg) decreased the hindlimb extensor phase of MES. The anticonvulsant effect of U50488H was attenuated by the general opioid antagonist, naloxone at a high dose, and by MR2266, a selective kappa antagonist, but not by naltrindole, a delta antagonist. Coadministration of gamma-aminobutyric acid (GABA)ergic drugs (diazepam, GABA, muscimol, and baclofen) and the N-methyl-D-aspartate (NMDA) receptor antagonist, dizocilpine (MK801), with U50488H augmented the anticonvulsant effect of the latter drug in mice. On the other hand, flumazenil, a central benzodiazepine (BZD) receptor antagonist, reversed the protective effect of diazepam and similarly, delta-aminovaleric acid (DAVA), a GABA(B) receptor antagonist, blocked the protective effect of baclofen, a GABA(B) agonist on the anti-MES action of U50488H. These BZD-GABAergic antagonists, namely, flumazenil or DAVA, on their own also counteracted the anti-electroshock seizure effect of U50488H given alone. However, mortality was not significantly altered in any of the above animal groups. Taken together, the findings have shown a possible role for multitude of important neurotransmitter systems, i.e., opioid (kappa), NMDA channel, GABA(A)-BZD-chloride channel complex, and GABA(B) receptors in the anticonvulsant action of U50488H.     

Cardiovascular responses to kappa opioid agonists in intact and adrenal demedullated rats

The effects of three kappa opioid agonists namely, bremazocine, tifluadom and U-50,488H were studied on blood pressure and heart rate in urethane-anesthetized normal and bilateral adrenal demedullated rats. Bremazocine (0.2, 0.4 and 0.6 mg/kg i.v.) produced a dose-dependent decrease in heart rate, while only 0.4 mg/kg bremazocine produced marked hypotension. The effect appeared to be long lasting because even at 60 min following drug administration the decreases in both heart rate and blood pressure continued. Bilateral adrenal demedullation did not change bremazocine-induced fall in blood pressure but the bradycardia was partially blocked. Tifluadom (0.1-0.4 mg/kg i.v.) produced an initial arrest of heart beat followed by bradycardia which recovered in about 60 min. Except for a very transient fall soon after drug administration no significant effect was observed on blood pressure. In adrenal demedullated rats, tifluadom induced initial arrest of heart was not affected but the subsequent bradycardia was blocked. U-50,488H (0.2, 0.4 and 0.6 mg/kg i.v.) produced dose-dependent bradycardia and hypotension both of which were blocked following bilateral adrenal demedullation. Naltrexone methylbromide (MRZ 2663 BR), a quaternary opioid antagonist, injected 5 min prior to U-50,488H, blocked its cardiovascular effects. The results suggest that kappa opioid agonists given i.v. depress cardiovascular system and these effects are mediated through the adrenal medulla and peripheral opioid receptors. The differential effects of kappa opioid agonists on blood pressure and heart rate suggest that either the three kappa agents interact differentially at the kappa opioid receptors or the subtypes of receptors for the kappa opioid exist.     

Withdrawal responses of guinea-pig isolated ileum following brief exposure to opiates and opioid peptides

Summary Withdrawal contractures following brief exposure of guinea-pig ileum to enkephalin analogues, dynorphin A-(1-13) and beta-endorphin and the opiate drugs morphine, ketocyclazocine, buprenorphine and MR2034 were investigated. Following 2 min contact with the ileum a withdrawal contracture was induced by washout of (Met⁵)- and (Leu⁵)-enkephalin and by several enkephalin analogues but not by washout of (D-Ala²,D-Leu⁵)-enkephalinamide or any of the other drugs tested. Addition of naloxone precipitated withdrawal to all opioids tested except the kappa receptor preferential drugs ketocyclazocine and MR2034 and the mu receptor partial agonist, buprenorphine.The heights of the withdrawal contractures to enkephalin analogues were found to be dependent on the concentration of agonist and of naloxone, and on the duration of the contact period of opioid with the ileum.The naloxone-precipitated withdrawal responses to morphine, dynorphin A-(1-13) and the washout withdrawal response to (Met⁵)-enkephalin were inhibited by substance P antagonists thus supporting the previous pro posal that substance P mediates the opiate withdrawal response.This study has shown that mu receptor agonists produced dependence in the guinea-pig ileum revealed by a withdrawal contracture on addition of naloxone, whereas dependence on kappa receptor agonists could not be revealed with naloxone. Since the guinea-pig ileum preparation has m and kappa receptors it was concluded that the endogenous opioid peptides, enkephalins, beta-endorphin and dynorphin A-(1-13), all induced dependence by acting on mu receptors in this preparation.     

Cardiovascular actions of the kappa-agonist, U-50,488H, in the absence and presence of opioid receptor blockade.

1. The cardiovascular actions of U-50,488H, a kappa-receptor agonist, were studied in rat isolated perfused hearts, and in anaesthetized rats, over concentrations or doses generally above those required to produce kappa-receptor-mediated effects. 2. U-50,488H dose-dependently decreased left-ventricular peak systolic pressure and beating rate in vitro and reduced blood pressure and heart rate in vivo. 3. Over the concentration range of 1-30 microM in vitro, and the dose-range of 0.5-32 mumol kg-1 in vivo, U-50,488H prolonged the P-R, QRS and Q-T intervals of the ECG. 4. The effects of U-50,488H were not antagonized by an opioid receptor antagonist, naloxone (1 microM or 8 mumol kg-1). Similarly, the opioid receptor antagonist, MR 2266, at 8 mumol kg-1 did not significantly reduce the cardiovascular actions of U-50,488H in vivo. 5. The actions of U-50,488H on responses to electrical stimulation were also studied. Over the dose range of 0.5-32 mumol kg-1, U-50,488H altered thresholds and effective refractory period. It had a biphasic action on thresholds for induction of ventricular fibrillation. Thresholds were decreased at lower doses (0.5-4 mumol kg-1) but increased at higher doses (8-32 mumol kg-1). The effects of lower doses were blocked by naloxone. Effective refractory period and threshold pulse width only increased with dose. 6. In conclusion, U-50,488H at high concentration, had direct depressant actions on cardiac contractility, electrical excitability and the ECG. These depressant effects were not antagonized by the opioid receptor antagonists, naloxone and MR 2266, and probably do not involve opioid receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kappa-opiates and urination: pharmacological evidence for an endogenous role of the kappa-opiate receptor in fluid and electrolyte balance

In prehydrated rats, the administration of kappa-opiate agonists such as bremazocine, ethylketocyclazocine or compound Upjohn-50,488 produced a dose-dependent increase in urine output and decreased the concentration of Na+ and K+ in the urine as compared to that of saline-treated rats. The diuretic effect of bremazocine lasted at least 3 h. The increase in urine output was independent of the hydration state of the rat since in non water-loaded animals, bremazocine produced proportionally as much diuresis and a decrease in the output of urine electrolytes of about the same magnitude as that observed in the prehydrated animals treated with the opioid. In contrast to the diuretic action of kappa-opiate agonists, the administration of antagonists with high affinity for the kappa-opiate receptor (Win 44,441 or Mr 2266) decreased dose dependently the output of urine and reduced very significantly the total output of Na+ and K+. Whereas 2 mg/kg naloxone did not block the bremazocine-induced urinary effects, 1 mg/kg Win 44,441 or Mr 2266 antagonized competitively the renal activity of bremazocine. The results are interpreted to suggest that the kappa-opiate receptor may be involved in the regulation of fluid and electrolyte balance.     

The specific opioid kappa-agonist U-50,488H inhibits low Km GTPase

DAGO, a specific opioid mu-agonist stimulated the low Km GTPase activity of membranes of the guinea-pig striatum at 1-1000 nM and this effect was antagonized by naloxone 10-100 nM. On the other hand, U-50,488H, a specific opioid kappa-agonist inhibited the low Km GTPase activity of membranes of the guinea-pig cerebellum at 1-100 nM. As this effect was antagonized by MR2266 but not by naloxone, the findings suggest that opioid mu- and kappa-receptor stimulation is probably linked to the stimulation and inhibition of low Km GTPase, respectively.     

Possible mechanism involved in the anticonvulsant action of butorphanol in mice

The study was designed to examine the effect of butorphanol, a classical opioid on convulsive behaviour using maximal electroshock (MES) test. An attempt was also made to investigate the role of possible receptor mechanisms involved. MES seizures were induced in mice via transauricular electrodes (60 mA, 0.2 s). Seizure severity was assessed by the duration of tonic hindlimb extensor phase and mortality due to convulsions. Intraperitoneal administration of butorphanol produced a dose-dependent (0.25-2 mg/kg) protection against hindlimb extensor phase. The anticonvulsant effect of butorphanol was antagonized by all the three opioid receptor antagonists (i.e., naloxone [mu], MR2266 [kappa], and naltrindole [delta], respectively). Coadministration of gamma-aminobutyric acid (GABA)-ergic drugs (diazepam, GABA, muscimol, and baclofen) and N-methyl-D-aspartate (NMDA) receptor antagonist, dizocilpine (MK801), with butorphanol augmented the anticonvulsant action of the latter drug. In contrast, flumazenil, a central benzodiazepine (BZD) receptor antagonist, reversed the facilitatory effect of diazepam on the anti-MES effect of butorphanol. Similarly, delta-aminovaleric acid (DAVA), a GABA(B) receptor antagonist, antagonized the facilitatory effect of baclofen, a GABA(B) agonist on anti-MES action of butorphanol. These BZD-GABAergic antagonists, flumazenil or DAVA, per se also counteracted the anti-MES effect of butorphanol given alone. These data exemplify the benefits of using the MES test, which is sensitive to opioidergic compounds and distinguished convulsive behavioural changes associated with GABAergic and NMDAergic effects. Taken together, the results implicate a role for multitude of neurotransmitter systems, i.e., opioid (mu, kappa, delta), NMDA channel, BZD-GABA(A) chloride channel complex, and GABA(B) receptors in the anti-MES action of butorphanol.     

Characterization of opioid receptors modulating the function of capsaicin-sensitive neurons in guinea-pig atria

Transmural nerve stimulation of isolated atria, obtained from reserpine-pretreated guinea-pigs, in the presence of atropine and the β₁-adrenoceptor-blocking drug CGP 20712A, induced a positive inotropic effect. [D-Ala², N-Me-Phe⁴, Gly⁵-ol] enkephalin (DAGO), [D-Ala², D-Leu⁵]enkephalin (DADLE), morphine and dynorphin dose dependently reduced the cardiac response to transmural nerve stimulation. The δ receptor selective agonist [D-Pen², D-Pen⁵]enkephalin (DPDPE), and the κ receptor agonist, U50488, were unable to modify the response. The inhibitory effect of all the active opioid agonists was antagonized by naloxone but not by the selective δ and κ opioid receptor antagonists, ICI 174.864 and MR 2266. These results suggest the presence on sensory nerve terminals of inhibitory opioid receptors belonging to the μ, but not to the δ and κ subtypes.     

Effects of pentazocine and other opioids on the potassium-evoked release of [3H]noradrenaline from guinea pig cortical slices

Noradrenaline release and its modulation via presynaptic opioid receptors were examined in guinea pig cortical slices. Slices preloaded with [3H]noradrenaline were superfused in the presence of desipramine (1 microM) and were stimulated by 16 mM K+. 1-Pentazocine inhibited the K+-evoked release of [3H]noradrenaline in a dose-dependent manner (3 x 10(-7)-10(-5) M), while d-pentazocine did not inhibit. This inhibitory effect of 1-pentazocine was antagonized by Mr 2266 (10(-6) M) but not by naloxone (10(-6) M). Among other opioids, dynorphin A-(1-13) and ethylketocyclazocine (kappa agonists) decreased the K+-evoked release of [3H]noradrenaline. Tyr-D-Ala-Gly-NMe-Phe-Gly-ol (DAGO, mu agonist) also inhibited [3H]noradrenaline release but was less potent than the kappa agonists. [D-Pen2,D-Pen5]enkephalin (DPDPE, delta agonist) and phencyclidine (sigma agonist) had no effects on the stimulated release of [3H]noradrenaline. Thus, it was shown that kappa receptors are the major subtype of opioid receptor involved in modulation of noradrenaline release in guinea pig cortex, and that 1-pentazocine inhibits the K+-evoked release of noradrenaline through activation of these receptors.     

Has the sun set on kappa3-opioid receptors?

Mu-opioid receptor agonists are a mainstay of clinical analgesia, despite the significant unwanted effects and dependence liability associated with drugs like morphine. The quest for opioids that produce analgesia with fewer undesirable effects has lead to the putative identification of multiple opioid receptor subtypes, despite the identification of only four opioid-related receptor genes. One such putative receptor subtype is the kappa3 receptor, activation of which supposedly produces analgesia in animals. In the present issue of this Journal, Olianas and co-workers have demonstrated that the prototypic kappa3 agonist naloxone benzoylhydrazone is actually a partial agonist at the cloned mu, delta, and kappa opioid receptors and an antagonist at opioid-like NOP receptors. Together with a recent study that showed that high-affinity naloxone benzoylhydrazone binding is abolished in triple mu/delta/kappa receptor knockout mice, the present study provides strong evidence that in vivo effects attributed to kappa3 receptor activation probably just reflect the combined actions of a particularly nonselective opioid drug. Indeed, molecular identification of any of the proposed subtypes of mu, delta, and kappa opioid receptors has proven elusive, suggesting that it is perhaps time to retire the notion of opioid receptor subtypes until definitive evidence for their existence is provided.     

Stereoselective enhancement of nociception by opioids in different strains of mice

The effects of Mr2096, Mr2097, diprenorphine, (-)bremazocine and Mr2266 on jumping latencies were evaluated in Swiss, CXBK, C57BL, CXBH and CBA strains of mice. Mr2096 and Mr2097 respectively produced analgesia and hyperalgesia in these strains of mice. Thus their effects on nociception were mediated by stereoselective opioid receptors. Diprenorphine also produced hyperalgesia which might be mediated by both mu and kappa receptors. Kappa receptors also appeared to mediate hyperalgesia at least in Swiss and CBA strains, as both (-)bremazocine and Mr2266 shortened the jumping latencies.