Diode-wing-pumped electro-optically Q-switched 2 µm laser with pulse energy scaling over ten millijoules.

Characteristics of diode-wing-pumped highly efficient Tm:LuAG lasers running both in continuous wave (CW) and electro-optical Q-switching regimes have been investigated. Using a simple plane-plane cavity, a maximum CW output power of 8.5 W has been achieved with a corresponding slope efficiency of 44.5% by "wing pumping" at 790 nm. With a V-shaped cavity, a diode-wing-pumped MgO:LiNbO3 crystal based electro-optically Q-switched Tm:LuAG laser at 2022.9 nm delivered a maximum pulse energy of 10.8 mJ and a minimum pulse width of 52 ns at a corresponding repetition rate of 100 Hz. To the best of our knowledge, the achieved CW output power and Q-switched pulse energy have both set records for all-solid-state Tm:LuAG lasers, which well reveals an efficient way to generate high-power and high-energy lasers at 2 µm wavelength.

Lead (Pb2+) modulation of potassium currents of guinea pig outer hair cells.

Outer hair cells (OHC) are mechanosensitive sensory cells of the inner ear cochlea and are involved in modulating the activity of inner hair cells in the transduction of an acoustic stimulus. Potassium (K(+)) currents play an important role in the sensory transduction process. K(+) currents were recorded from acutely dissociated OHC obtained from the guinea pig organ of Corti. The whole-cell patch clamp technique was employed. We identified a channel that exhibited outward current of the delayed rectifier type (Kv). Kv channels mediating inward currents carried by potassium ions were also identified and took on the appearance of a previously described inwardly rectifying current. Lead (Pb(2+)) acetate at concentrations of 0.1, 1.0, 10, and 100 microM was bath applied. Time to activation for outward-going current was not affected by Pb(2+). The time course of Pb(2+) effects was seen as a dose-dependent reduction of K(+) current over time, with very little or no recovery after washout. Pb(2+) inhibited the outward Kv relative current with values of 0.10, 0.14, 0.18, and 0.30 at 0.1, 1.0, 10, and 100 microM, respectively. Pb(2+) did not modulate time to activation, peak current, or inactivation of inward I(K). The effects of Pb(2+) on the potassium currents of OHC are not remarkable and therefore OHC are probably not a major cause of purported peripheral hearing loss observed in Pb(2+)-exposed animals and humans.

Effects of acute administration of L-arginine on morphine antinociception and morphine distribution in central and peripheral tissues of mice.

The effect of acute treatment with L-arginine, a substrate for nitric oxide synthase (NOS) that forms NO, an important second messenger, on morphine antinociception and distribution of morphine in central and peripheral tissues of male Swiss-Webster mice was determined. The antinociception activity of morphine (10 mg/kg, s.c.) was attenuated by 400 and 800 mg/kg doses of L-arginine, but a lower dose (200 mg/kg) had no effect. D-Arginine (200-800 mg/kg) did not modify morphine antinociception. The dose of L-arginine (200 mg/kg) that did not modify morphine antinociception also did not alter the distribution of morphine in brain regions and spinal cord. A dose of 800 mg/kg of L-arginine produced a significant decrease in the concentration of morphine in midbrain and spinal cord. The highest dose of L-arginine (800 mg/kg) also increased the concentration of morphine in spleen. None of the doses of L-arginine modified the concentration of morphine in serum or urine. The results suggests that acute activation of the NO system attenuates morphine antinociception possibly by inhibiting its uptake in central sites (midbrain and spinal cord) involved in antinociceptive actions.

Effect of chronic administration of L-arginine, NG-nitro-L-arginine or their combination on morphine concentration in peripheral tissues and urine of the mouse.

1. Chronic administration of L-arginine (200 mg/kg, i.p) twice a day for 4 days decreased the antinociceptive response to subcutaneously, but not to intracerebroventricularly, administered morphine in male Swiss-Webster mice, as measured by the tail-flick test. 2. The decreased antinociceptive response to morphine was reversed by concurrent administration of NG-nitro-L-arginine (L-NNA) (5 mg/kg, IP), an inhibitor of nitric oxide synthase. 3. The concentrations of morphine in mice treated chronically with L-arginine and then given morphine (10 mg/kg, SC) were determined in the peripheral tissues. L-Arginine treatment significantly increased the concentration of morphine in spleen and lungs, did not modify it in liver, kidneys and urine. L-NNA by itself had no effect on the distribution of morphine in peripheral tissues but reversed the changes induced by chronic treatment with L-arginine. 4. Acute administration of L-arginine (200 mg/kg, IP) did not modify either the morphine antinociception or the morphine distribution in peripheral tissues. 5. Previous studies from this laboratory indicated that chronic treatment with L-arginine decreases the concentration of morphine in several brain regions and spinal cord of mice. 6. The facts that chronic treatment with L-arginine does not alter antinociception induced by ICV administered morphine and it increases the concentration of morphine in peripheral tissues while decreasing it in brain regions after peripheral administration of morphine suggest that the decreased antinociception induced by subcutaneously administered morphine may be related to its decreased entry into the brain.

Evidence for a role of N-methyl-D-aspartate receptors in L-arginine-induced attenuation of morphine antinociception.

Twice daily injections of L-arginine (50, 100 or 200 mg/kg, i.p.) for 4 days dose-dependently, decreased morphine antinociception in male Swiss-Webster mice as measured by the tail-flick test. To determine the possible role of N-methyl-D-aspartate (NMDA) receptor in the action of L-arginine, the effects of MK-801, a noncompetitive antagonist of the NMDA receptor and of LY 235959, a competitive antagonist of the NMDA receptor on L-arginine-induced attenuation of morphine antinociception were determined. MK-801 (0.01-0.10 mg/kg, i.p.) or LY 235959 (1.0-4.0 mg/kg, i.p.) given 10 min before each injection of L-arginine (200 mg/kg, i.p.) reversed the action of the letter in a dose-dependent manner on morphine antinociception. It is concluded that NMDA receptors are involved in the action of L-arginine in attenuating morphine antinociception.

Effects of acute and chronic administration of L-arginine on the antinociceptive action of morphine-6-beta-D-glucuronide.

Chronic administration of L-arginine (200 mg/kg i.p.) but not of D-arginine (200 mg/kg i.p.) twice a day for 4 days decreased the antinociceptive response to subcutaneously administered morphine-6-beta-D-glucuronide (M6G), a potent metabolite of morphine, in male Swiss-Webster mice as measured by the tail-flick test. However, the antinociceptive response to intracerebroventricularly administered M6G was unaffected by chronic treatment with L-arginine. The decreased antinociceptive response to M6G (s.c.) was reversed by concurrent administration of NG-nitro-L-arginine (5 mg/kg i.p.), an inhibitor of nitric oxide synthase. Acute administration of L-arginine (200 mg/kg i.p.) had no effect on M6G-induced antinociception, but higher doses (400 and 800 mg/kg i.p.) decreased it. Since the antinociceptive response to centrally administered M6G was unaffected by chronic L-arginine treatment, the decreased antinociceptive response of peripherally administered M6G may be related to a decrease of M6G entry into brain structures responsible for antinociceptive action.

Effects of NMDA receptor antagonists on delta1- and delta2-opioid receptor agonists-induced changes in the mouse brain [3H]DPDPE binding.

Male Swiss-Webster mice were injected intracerebroventricularly (i.c.v.) with [D-Pen2,D-Pen5]enkephalin (20 microg/mouse) twice a day for 2 days. This procedure resulted in down-regulation of binding sites for [3H][D-Pen2,D-Pen5]enkephalin as evidenced by a 52% decrease in the Bmax value. Twice daily injections of (+)-5-methyl-10,11-dihydro-5H-dibenzo-[a,d]-cyclohepten-5,10-imine (MK-801) (0.1 mg/kg, i.p.) or [(-)3-SR,4a-RS,8a-SR-6-(phosphonomethyl)-1,2,3,4,4a,5,6,7,8,8a-decahy droisoquinoline-3-carboxylic acid] (LY 235959) (2 mg/kg, i.p.), the noncompetitive and competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor, respectively, for 2 days did not alter the Bmax or Kd value of [3H][D-Pen2,D-Pen5]enkephalin binding to the mouse brain. Concurrent treatment of MK-801, but not of LY 235959 with [D-Pen2,D-Pen5]enkephalin, reversed the decreases in Bmax value of [3H][D-Pen2,D-Pen5]enkephalin. Twice daily injections of [D-Ala2,Glu4] deltorphin II (20 microg/mouse) for 2 days caused an increase in the Kd value, but not the Bmax value of [3H][D-Pen2,D-Pen5]enkephalin to bind to brain membranes. Concurrent treatment of [D-Ala2,Glu4]deltorphin II with LY 235959 reversed the increase in Kd value of [3H][D-Pen2,D-Pen5]enkephalin binding induced by multiple injections of [D-Ala2,Glu4]deltorphin II, but MK-801 had no effect. The results suggest that multiple injections of delta1- and delta2-opioid receptor agonists down-regulate delta1-opioid receptors of the brain by modifying Bmax and Kd values of [3H][D-Pen2,D-Pen5]enkephalin binding, respectively. MK-801 and LY 235959 reverse delta1- and delta2-opioid receptor agonists-induced down-regulation of brain delta1-opioid receptor, respectively, apparently by different mechanisms. It is concluded that short term treatment of mice with delta1-opioid receptor agonist down-regulates brain delta1-opioid receptors by decreasing Bmax of the ligand which is partially reversed by concurrent treatment with MK-801 but not by LY 235959. On the other hand, short term treatment of mice with delta2-opioid receptor agonist down-regulates brain delta1-opioid receptors by increasing Kd of the ligand which is partially reversed by concurrent treatment with LY 235959 but not by MK-801.

Mechanism of attenuation of morphine antinociception by chronic treatment with L-arginine.

The effects of twice-daily injections of L-arginine or D-arginine (200 mg/kg i.p.) for 4 days on morphine-induced antinociception, brain nitric oxide synthase activity and brain and serum distribution of morphine and brain mu-opioid receptors labeled with [3H][D-Ala2,MePhe4,Gly5-ol]enkephalin were determined in male Swiss-Webster mice. Chronic treatment with L-arginine, but not D-arginine, decreased the antinociceptive response to morphine in mice, increased the activity of nitric oxide synthase in the midbrain and decreased brain levels of morphine, compared with vehicle-injected controls. Significant decreases in morphine levels were observed in midbrain, pons and medulla, hippocampus, striatum and spinal cord of L-arginine-treated mice, in comparison with vehicle-injected mice. However, the levels of morphine in cortex, amygdala and hypothalamus of L-arginine- or D-arginine-treated mice did not differ from those of vehicle-injected controls. Acute treatment with L-arginine (200 mg/kg i.p.) or D-arginine (200 mg/kg i.p.) did not modify either morphine antinociception or morphine distribution in brain regions or the spinal cord. Chronic administration of L-arginine or D-arginine did not alter the Bmax or Kd values of [3H][D-Ala2,MePhe4,Gly5-ol]enkephalin binding to the mouse brain membranes. These results suggest that chronic treatment with L-arginine reduces the antinociceptive effect of morphine by increasing brain nitric oxide synthase activity and by decreasing the concentration of morphine in certain brain regions and spinal cord.

N(G)-nitro-L-arginine reverses L-arginine induced changes in morphine antinociception and distribution of morphine in brain regions and spinal cord of the mouse.

Twice daily injections of L-arginine (200 mg/kg, i.p.), a precursor for nitric oxide (NO), for 4 days decreased morphine antinociception in male Swiss-Webster mice. Chronic treatment with L-arginine also produced significant decreases in morphine levels in midbrain, pons and medulla, hippocampus, corpus striatum and spinal cord of mice following an injection of morphine (10 mg/kg, s.c.) in comparison to vehicle-injected mice. N(G)-nitro-L-arginine (L-NNA), an inhibitor of NO synthase (NOS), (5 mg/kg, i.p.) given prior to each injection of L-arginine reversed the effects of the latter on morphine antinociception and decreases in morphine levels in brain regions and spinal cord. Chronic injections of L-NNA alone did not modify either morphine antinociception or morphine distribution in brain regions and spinal cord of mice. These results suggest that decreases in morphine antinociception by chronic treatment with L-arginine is related to the decreases in the entry of morphine in the central sites. The reversal of L-arginine-induced effects by L-NNA suggests that NO-NOS system may be playing a critical role in the regulation of blood-brain barrier to morphine.

Effects of some 7-arylidene and 7-heteroarylidene morphinan-6-ones on the antinociceptive activity of [D-Pen2, D-Pen5]enkephalin and [D-Ala2, Glu4]deltorphin II and on multiple opioid receptors.

The in vivo and functional effects of several 7-arylidene and 7-heteroarylidene morphinan-6-ones were determined at the mu-, delta-, and kappa-opioid receptors using the guinea pig brain membranes, guinea pig ileum (GPI), and mouse vas deferens (MVD). In vivo effects included the antagonism by these compounds given subcutaneously on the antinociceptive actions of intracerebroventricularly injected [D-Pen2, D-Pen5]enkephalin (DPDPE) and [D-Ala2, Glu4]deltorphin II (deltorphin II), the highly selective putative delta 1- and delta 2-opioid receptor agonists. Finally, the partition coefficients of these compounds were estimated (CLOGP) and determined experimentally at pH 7.4 in the 1-octanol/water system. Compared with E-7-benzylidenenaltrexone (BNTX), most compounds except for E-7-(4-chlorobenzylidene)naltrexone, were more potent at delta-opioid receptors than at the mu-opioid receptor, whereas, in comparison to the kappa-opioid receptor, the activities of the E-7-arylidene or E-7-heteroarylidene naltrexone derivatives at the delta-receptor were in the following order, where the 7-substituents were: 4-fluorobenzylidene- > benzylidene > 3-pyridylmethylene- > 4-pyridylmethylene- > 1-methyl-2-imidazolylmethylene > 4-chlorobenzylidene. In the MVD preparation, the potencies at the delta-opioid receptor, in comparison to BNTX, were in the following order, where the 7-substituents were: benzylidene = 1-methyl-2-imidazolylmethylene- > 4-fluorobenzylidene- = 3-pyridylmethylene- = 4-pyridylmethylene-. All compounds antagonized delta 1, and delta 2-opioid receptor agonist-induced analgesia. The antagonist potencies at the delta 1-opioid receptor were in the following order, where the 7-substituents were: benzylidene- > 4-chlorobenzylidene- > 4-fluorobenzylidene- > 3-pyridylmethylene- > 1-methyl-2-imidazolymethylene- approximately 4-pyridylmethylene-, whereas at the delta 2-opioid receptor, the order was benzylidene- > 4-chlorobenzylidene- > 4-fluorobenzylidene- > 3-pyridylmethylene- > 1-methyl-2-imidazolymethylene- > 4-pyridylmethylene. In general, all compounds exhibited greater potency at the delta 2- than delta 1-opioid receptor. The computed partition coefficients were, as expected, greater than the apparent log P values, which were determined experimentally. Generally, the lipophilicity values in decreasing order were: 4-chlorobenzylidene- > 4-fluorobenzylidene- > benzylidene > 3-pyridylmethylene- = 4-pyridylmethylene- > 1-methyl-2-imidazolylmethylene-. In general, the benzylidene and 4-pyridylmethylene derivatives, which have medium lipophilicities, were equally effective at the delta 1- and delta 2-receptors; the 3-pyridylmethylene and 1-methyl-2-imidazolylmethylene derivatives had lower lipophilicities and were more selective for the delta 2- than delta 1-receptor; the 4-chlorobenzylidene and 4-fluorobenzylidene derivatives were more lipophilic and had intermediate activity. The plot of pED50 values for the in vivo tests for the delta 1- and delta 2-receptors showed that the two receptors are not independent with respect to this series of compounds.

Up-regulation of brain N-methyl-D-aspartate receptors following multiple intracerebroventricular injections of [D-Pen2, D-Pen5] enkephalin and [D-Ala2, Glu4]deltorphin II in mice.

The effects of chronic administration of [D-Pen2, D-Pen5]enkephalin and [D-Ala2, Glu4]deltorphin II, the selective agonists of the delta 1- and delta 2-opioid receptors, on the binding of [3H]MK-801, a noncompetitive antagonist of the N-methyl-D-aspartate receptor, were determined in several brain regions of the mouse. Male Swiss-Webster mice were injected intracerebroventricularly (i.c.v.) with [D-Pen2, D-Pen5]enkephalin or [D-Ala2, Glu4]deltorphin II (20 micrograms/mouse) twice a day for 4 days. Vehicle injected mice served as controls. Previously we have shown that the above treatment results in the development of tolerance to their analgesic activity. The binding of [3H]MK-801 was determined in brain regions (cortex, midbrain, pons and medulla, hippocampus, striatum, hypothalamus and amygdala). At 5 nM-concentration, the binding of [3H]MK-801 was increased in cerebral cortex, hippocampus, and pons and medulla of [D-Pen2, D-Pen5]enkephalin treated mice. In [D-Ala2, Glu4]deltorphin II treated mice, the binding of [3H]MK-801 was increased in cerebral cortex and hippocampus. The changes in the binding were due to increases in the Bmax value of [3H]MK-801. It is concluded that tolerance to delta 1- and delta 2-opioid receptor agonists is associated with up-regulation of brain N-methyl-D-aspartate receptors, however, some brain areas affected differ with the two treatments. The results are consistent with the recent observation from this laboratory that N-methyl-D-aspartate receptors antagonists block tolerance to the analgesic action of delta 1- and delta 2-opioid receptor agonists.

[Frequency dependence of substance P release by electroacupuncture in rat spinal cord].

Previous studies in our laboratory have shown that electroacupuncture (EA) using different frequencies produced differential opioid peptides release in the spinal cord of rats and human beings. In the present study we observed the frequency dependence of substance P (SP) release from rat spinal cord, with the frequencies of EA set at 2, 4, 8, 15, 30 and 100 Hz. The spinal perfusate was collected in 3 periods of 30 min before, during and after EA, and the immunoreactive SP (SP-ir) was measured by radioimmunoassay (RIA). The effectiveness of EA-induced analgesia was assessed by tail flick latency (TFL). Rats showing an increase of TFL over 40% was considered as EA responder. The results showed that in the responders, SP-ir in spinal perfusate showed a moderate decrease during 2 Hz EA, (P < 0.01 compared with baseline level), no change in the 4 Hz EA group, and a marked increase during 8, 15, 30 and 100 Hz EA (P < 0.01), with maximal increase occurring at 15 Hz (P < 0.001). The above results suggest that EA may induce upward or downward modulation in SP-ir release depending on the frequency of EA. However in the non-responder rats no change in spinal fluid SP-ir content was observed. This suggests that changes in SP-ir release have same causal relation with the analgesia induced by EA stimulation.

Effects of morphine-3-glucuronide on the antinociceptive activity of peptide and nonpeptide opioid receptor agonists in mice.

The effects of morphine-3-glucuronide (M3G), a metabolite of morphine, were determined on the antinociceptive actions, as measured by the tail flick test, of morphine, a mu-opioid receptor agonist, of U-50,488H, a kappa-opioid receptor agonist of [D-Pen2, D-Pen3]enkephalin (DPDPE), a delta 1-opioid receptor agonist, and of [D-Ala2,Glu4]deltorphin II (deltorphin II), a delta 2-opioid receptor agonist in mice. Morphine administered ICV (2.5 micrograms/ mouse) or SC (10 mg/kg), U-50,488H (25 mg/kg, IP), DPDPE (15 micrograms/mouse; ICV), and deltorphin II (15 micrograms/mouse, ICV) produced antinociception in mice. Intraperitoneal or ICV injections of M3G did not produce any effect on the tail flick latency nor did it affect the antinociception-induced by morphine, U-50,488H, DPDPE, or deltorphin II. Previously M3G has been shown to antagonize the antinociceptive effects of morphine in the rat. It is concluded that in the mouse, M3G neither produces hyperalgesia nor modifies the actions of mu-, kappa-, delta 1-, or delta 2-opioid receptor agonists.

[Study on a role of substance P in the spinal mechanisms of electroacupuncture analgesia].

It was found in the present study that low frequency (2Hz) electroacupuncture (EA) stimulation caused a decrease of the content of substance P immunoreactivity (SP-ir), whereas medium-(15Hz), high-(100Hz) and dense-disperse (D-D)- (2/15Hz) frequencies EA stimulation induced an increase of the content of SP-ir in the rat spinal fluid. EA analgesia induced by medium-, high- or D-D mode frequency was suppressed by nonpeptide SP (NK1) receptor antagonists CP96345 or RP67580 administered intrathecally (i.t.). Both the attenuation of SP release by low frequency EA and the potentiation of SP release by medium frequency EA in the spinal cord were blocked by the opioid receptor antagonist naloxone (i.t.). These results suggest that a decrease of release of SP-ir by low frequency and an increase of it by medium-, high- and D-D mode frequencies in the spinal cord facilitate analgesia.

Reversal of electroacupuncture tolerance by CCK-8 antiserum: an electrophysiological study on pain-related neurons in nucleus parafascicularis of the rat.

Two varieties of neurons were found in nucleus parafascicularis (pf) of the rat: one responds to noxious stimuli with an increase in firing (pain-excited neuron, PEN), the other with a decrease in firing (pain-inhibited neuron, PIN). Electroacupuncture (EA) has been shown to suppress PEN and excite PIN, which can be taken as an electrophysiological index for EA analgesia. This effect of EA subsided after prolonged (6 h) EA stimulation, suggesting the development of tolerance to EA. Intracerebroventricular (icv) injection of CCK-8 antiserum aiming at neutralizing endogenously released CCK-8 resulted in a complete restoration of the EA effect. Normal rabbit serum was not effective. CCK-8 antiserum per se did not affect the firing pattern of the PEN or PIN in nontolerant rat. The results obtained from single neuron recording in anesthetized animals thus confirmed those obtained in intact animals using the tail flick as the end point, implying that an excess of endogenously released CCK-8 may constitute one of the mechanisms for the development of EA tolerance.