Phase transformation of plasma-sprayed hydroxyapatite coating with preferred crystalline orientation.

Highly oriented hydroxyapatite coatings (HACs) were obtained on titanium substrates through a radio-frequency thermal plasma spraying (TPS) method. XRD patterns showed that the HACs had crystallites with [001] preferred orientation vertical to the coating's surface. XRD results also indicated that tetracalcium phosphate crystallites in the as-sprayed HAC were oriented in the (100) direction. XRD peaks corresponding to tetracalcium phosphate, tricalcium phosphate and calcium oxide were absent after heat and hydrothermal treatment. The orientation degree of the HAC was influenced little by such post-heat treatments. Considering the crystallographic relationship between the tetracalcium phosphate in the as-sprayed HAC and the HA crystallites formed in the heat-treated HAC, these XRD results indicate that the tetracalcium phosphate in the as-prepared coatings transformed topotaxially into HA during the post-heat treatment. TEM and SEM analyses of the highly oriented HAC were conducted. The characteristic lamellar structure of TPS deposits was observed in cross-sections of the HAC. A prismatic texture was also observed in magnified SEM images. TEM observation showed that 200-800-nm-wide prismatic crystallites were formed in HA splats, and their longitudinal axis was oriented vertically to the coating's surface. SAD patterns showed that the longitudinal axis of the prismatic crystallites corresponded to the [001] zone axis of the HA crystal.

ROM7/BEM4 encodes a novel protein that interacts with the Rho1p small GTP-binding protein in Saccharomyces cerevisiae.

The RHO1 gene encodes a homolog of the mammalian RhoA small GTP-binding protein in the yeast Saccharomyces cerevisiae. Rho1p is localized at the growth site and is required for bud formation. The RHO1(G22S, D125N) mutation is a temperature-sensitive and dominant negative mutation of RHO1, and a multicopy suppressor of RHO1(G22S, D125N), ROM7, was isolated. Nucleotide sequencing of ROM7 revealed that it is identical to the BEM4 gene (GenBank accession number L27816), although its physiological function has not yet been reported. Disruption of BEM4 resulted in the cold- and temperature-sensitive growth phenotypes, and cells of the deltabem4 mutant showed abnormal morphology, suggesting that BEM4 is involved in the budding process. The temperature-sensitive growth phenotype was suppressed by overexpression of RHO1, ROM2, which encodes a Rho1p-specific GDP/GTP exchange factor, or PKC1, which encodes a target of Rho1p. Moreover, glucan synthase activity, which is activated by Rho1p, was significantly reduced in the deltabem4 mutant. Two-hybrid and biochemical experiments revealed that Bem4p directly interacts with the nucleotide-free form of Rho1p and, to lesser extents, with the GDP- and GTP-bound forms of Rho1p, although Bem4p showed neither GDP/GTP exchange factor, GDP dissociation inhibitor, nor GTPase-activating protein activity toward Rho1p. These results indicate that Bem4p is a novel protein directly interacting with Rho1p and is involved in the RHO1-mediated signaling pathway.

[Unusual total anomalous pulmonary venous connection; report of a case].

We report a case of a 7-hour-old infant with total anomalous pulmonary venous connection having abberant origin of the vertical vein. His clinical condition presented hemodynamically severe pulmonary vein obstruction as common pulmonary vein atresia. Without making the definitive diagnosis, he underwent the operation at 11 hours after birth under cardiopulmonary bypass. During the operation, we could not identify the drainage vein. Although we ligated the vessel sized less than 2 mm in diameter draining into superior vena cava which was suspected to be the vertical vein. After the operation pulmonary edema was severe, but his clinical condition improved by using nitric oxide without extracorporeal membrane oxygenation (ECMO) support. Postoperative cardiac catheterization showed normal cardiac function without pulmonary venous obstruction, and moreover identified the drainage vein originated from right upper pulmonary vein. To the best of our knowledge, this is the first case where the vertical vein originated from right upper pulmonary vein. Immediate surgical treatment before the circuratory exacerbation improve the outcome for this congenital anomaly.

Translocation of activated Rho from the cytoplasm to membrane ruffling area, cell-cell adhesion sites and cleavage furrows.

Rho small GTP-binding protein regulates various cell functions, such as formation of stress fibers and focal adhesions, cell motility, membrane ruffling, cytokinesis and smooth muscle contraction in mammalian cells and bud formation in the yeast Saccharomyces cerevisiae. As to the functioning sites of Rho in Saccharomyces cerevisiae, we have recently shown that RHO1 protein, a homologue of mammalian RhoA, is concentrated to the growth region of the cells where cortical actin patches are clustered. However, in mammalian cells, the functioning sites of Rho have not yet been studied. In the present study, MDCK cell lines stably expressing myc-tagged RhoA (myc-RhoA) were prepared and localization of myc-RhoA was first immunohistochemically examined using an anti-myc antibody. In the resting cells, almost all of myc-RhoA was observed in the cytosol. When the cells were stimulated with phorbol ester or hepatocyte growth factor, membrane rufflings were induced and myc-RhoA was translocated to the membrane ruffling area. Moreover, myc-RhoA was translocated from the cytosol to the cell-cell adhesion sites when the cells were transferred from a low to normal Ca2+ medium. RhoA was also concentrated to the cleavage furrows during cytokinesis in Swiss 3T3 cells. Translocation of myc-RhoA to the membrane ruffling area was inhibited by prior microinjection into the cells of Rho GDI, a negative regulator of Rho which inhibits activation of Rho, or of C3, an exoenzyme of Clostridium botulinum which ADP-ribosylates Rho and inhibits its functions, indicating that both activation and functioning of Rho are essential for the translocation of Rho. The ERM (Ezrin, Radixin, Moesin) family members were colocalized with RhoA at all of these sites. However, RhoA was not apparently observed at the focal adhesion plaque where vinculin was localized. These results suggest that at least one of the functioning sites of Rho is the ERM family-controlled actin filament/plasma membrane association sites.

Stereochemical structure recognized by the L-fucose-specific hemagglutinin produced by Streptomyces sp.

A hemagglutinin has been purified 4000-fold from the culture filtrate of a strain of Streptomyces by affinity chromatography. The purified preparation was judged to be homogeneous by gel electrophoresis and its molecular weight was estimated to be about 70 000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis. It may exhibit its full hemagglutinating activity in the monomer form. This hemagglutinin strongly agglutinated human blood group O erythrocytes and was inhibited by L-fucose. It was, however, distinct from the known L-fucose-specific hemagglutinins; first, the hemagglutinating activity of the purified preparation was more than 100-times stronger than that of others; second, D-mannose was a potent inhibitor of this hemagglutinin besides L-fucose but not or scarcely inhibitory to others; and third, p-nitrophenyl-beta-L-fucoside was more inhibitory to this hemagglutinin than p-nitrophenyl-alpha-L-fucoside as opposed to the case of others.

Respiratory modulation of muscle sympathetic nerve activity in patients with chronic heart failure.

BACKGROUND: Sympathoexcitation and respiratory instability are closely related to worsening of chronic heart failure. To elucidate the dynamic nature of respiratory modulation of sympathetic activity in patients with heart failure, we studied within-breath variation of muscle sympathetic nerve activity (MSNA) under various ventilatory volumes. METHODS AND RESULTS: MSNA, blood pressure, and respiratory flow were recorded in 23 patients with left ventricular ejection fraction

Ventricular load optimization by unloading therapy in patients with heart failure.

The effects of unloading a depressed heart were assessed in terms of optimal coupling between the ventricle and arterial system. To assess the effects of preload on ventricular load coupling, preload was reduced with a lower body negative pressure of -20 mm Hg. Nitroprusside was used to evaluate the effects of afterload on the coupling under the condition that preload reduction was comparable to that with lower body negative pressure. In 13 patients with heart failure (ejection fraction 32 +/- 3%, mean +/- SE), direct arterial pressure was simultaneously recorded with the left ventricular echocardiogram as the pressure was elevated by phenylephrine. Left ventricular contractile properties were defined by the slope (Ees) of the end-systolic pressure-volume relation. The effective arterial elastance (Ea) was expressed by the slope of the end-systolic pressure-stroke volume relation. Left ventricular external work, end-systolic potential energy and work efficiency, defined as external work per pressure volume area (external work + potential energy), were determined. Baseline ventricular load coupling in these patients was characterized by an increase in the ratio of arterial elastance to ventricular elastance (Ea/Ees) (1.96 +/- 0.31). This ratio decreased significantly, to 1.45 +/- 0.22, with nitroprusside, and increased to 2.37 +/- 0.34 with lower body negative pressure. Therefore, end-systolic potential energy was decreased by nitroprusside but was unaltered by lower body negative pressure while external work was comparably decreased by both manipulations, indicating that work efficiency was significantly augmented with nitroprusside but declined with lower body negative pressure.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute hemodynamic effects of a new inotropic agent (OPC-8212) in patients with congestive heart failure.

The acute effects of OPC-8212, a newly synthesized orally effective inotropic agent, were assessed clinically. Eleven patients with moderate congestive heart failure received a single mean dose of 6.5 mg/kg body weight of the drug. Eight hours after administration, the cardiac and stroke work indexes increased by 11% (p less than 0.01) and 20% (p less than 0.005), respectively, with concomitant decreases in the diastolic pulmonary artery (25%, p less than 0.005) and right atrial pressures (33%, p less than 0.01). There were no significant changes in blood pressure or heart rate. The contractile state of the left ventricle was also assessed by the shift of the Starling curve. To construct the function curve, lower body negative pressure was used to regulate the venous return to the heart. An inotropic effect of the agent was confirmed by the shift of this function curve upward and to the left, even when an augmentation of the cardiac output was masked by the marked reduction in preload. The hemodynamic and clinical effects of OPC-8212 were encouraging and the drug appears to be promising for the treatment of congestive heart failure.

Altered inotropic and lusitropic responses to heart rate in conscious dogs with tachycardia-induced heart failure.

OBJECTIVES: The effects of increasing heart rate on left ventricular contraction and relaxation were examined in conscious dogs with tachycardia-induced heart failure under autonomically blocked conditions. BACKGROUND: Previous studies using isolated myocardium have shown attenuated positive inotropic responses to stimulation frequency in heart failure. However, these responses have not been well examined in intact preparations in the presence of heart failure with autonomic system blockade, where the intrinsic ventricular responses to increasing heart rate could be revealed. METHODS: Seven dogs were instrumented with a micromanometer and a conductance volume catheter. After autonomic blockade to eliminate neural reflexes, left ventricular contractile properties were quantified by the slope of the end-systolic pressure-volume relation (ventricular elastance), and left ventricular relaxation was assessed by the time constant of isovolumetric ventricular pressure decay. RESULTS: Increasing the heart rate by 60 beats/min enhanced ventricular elastance by 71 +/- 18% (mean +/- SD) and decreased end-systolic volume by 6 +/- 5% in normal hearts. In failing hearts, ventricular elastance increased by only 21 +/- 20%, and end-systolic volume did not change appreciably. Although the reduction in left ventricular end-diastolic and minimal pressures by tachycardia was smaller in the failing heart, ventricular relaxation rate remained unaltered both in the normal heart and in the failing heart. CONCLUSIONS: Under conscious but autonomically blocked conditions, effects of increasing heart rate on the failing left ventricle are characterized by a predominant attenuation of the inotropic response rather than of the lusitropic response.

Modulation of left ventricular diastolic distensibility by collateral flow recruitment during balloon coronary occlusion.

OBJECTIVES: The goals of this study were to elucidate the scaffolding effect of blood-filled coronary vasculature and to determine the functional role of recruited collateral flow in modulating left ventricular (LV) distensibility during balloon coronary occlusion (BCO). BACKGROUND: Although LV distensibility is an important factor affecting acute dilation after myocardial infarction, the response of LV diastolic pressure-volume (P-V) relations to coronary occlusion is inconsistent in humans. METHODS: Micromanometer and conductance derived LV P-V loops were serially obtained from 16 patients undergoing percutaneous transluminal coronary angioplasty. Coronary collateral flow recruitment was angiographically evaluated by contralateral and ipsilateral contrast injection during BCO. RESULTS: In the group with poor collateral flow (grades 0-I; n = 8), BCO resulted in a downward and rightward shift of the diastolic P-V relations, where end-diastolic volume (EDV) increased by 13% (p < 0.05) without appreciable change in end-diastolic pressure (EDP; 18 +/- 6 to 18 +/- 8 mm Hg). In contrast, BCO in the group with good collateral flow (grades II-III; n = 8) shifted the diastolic P-V relations upward to the right with a concomitant increase in minimal pressure (min-P; 6 +/- 4 to 10 +/- 5 mm Hg, p < 0.05), EDP (15 +/- 7 to 21 +/- 9 mm Hg, p < 0.05) and EDV (+/- 10%, p < 0.05). Reactive hyperemia after balloon deflation caused a rapid and parallel upward shift of the diastolic P-V relations with a marked increase in min-P and EDP, especially in the group with poor collateral flow, before any improvement in LV contraction or relaxation abnormalities. CONCLUSIONS: Grades of coronary filling, either retrograde or anterograde, abruptly modulate LV distensibility through the rapid scaffolding effect of coronary vascular turgor.

Time-varying spectral analysis of heart rate and left ventricular pressure variability during balloon coronary occlusion in humans: a sympathoexicitatory response to myocardial ischemia.

OBJECTIVES: We assessed time-varying spectral components of heart rate and left ventricular (LV) pressure variability during coronary angioplasty to elucidate dynamic autonomic responses to transient myocardial ischemia. BACKGROUND: Sympathoexcitatory reflexes elicited by acute coronary occlusion are rarely addressed in the clinical settings because of a lack of technique to monitor transient changes in sympathetic activation. METHODS: RR interval and LV pressure and volume were serially recorded in 14 patients with effort angina during balloon coronary angioplasty. Wavelet analysis was applied for determination of nonstationary spectral components of RR interval and LV peak pressure variability. RESULTS: The wavelet analysis revealed that coronary occlusion provoked low-frequency (LF) fluctuations of RR interval (seven patients) and LV peak pressure (six patients) at 0.06 +/- 0.01 Hz, but not in the remaining patients. Following the balloon inflation, the LF component of RR interval began to increase after the onset of myocardial ischemia, peaked at about 80 s, and then declined in the late phase of inflation. Consequently, the ratio of low to high frequency component rose to be significantly greater in the LF augmentation group than in the no LF augmentation group in the middle phase of coronary occlusion. The patients with no LF augmentation had little evidence of myocardial ischemia as reflected by changes in ST segment and LV systolic function during coronary occlusion. CONCLUSIONS: The wavelet analysis of RR interval and LV pressure variability clearly showed a dynamic profile of spectral components in response to transient coronary artery occlusion. The resultant regional myocardial ischemia elicited a profound sympathoexcitatory response followed by a gradual suppression. This method provides a useful tool to gain a new insight into the nonstationary autonomic influence on the cardiovascular system.

Pathogenesis and prevention of HTLV-1-associated diseases.

HTLV-1 is an important factor involved in various diseases including adult T-cell leukemia/lymphoma and HTLV-1 associated myelopathy/tropical spastic paraparesis. Amount of HTLV-1 provirus integrated in human peripheral blood mononuclear cells might be a candidate for a risk factor in the manifestation of HTLV-1 associated diseases. Experimental animal models would be useful to dissect the pathogenesis of HTLV-1 associated diseases. We present rat and mouse models of HTLV-1 infection. Using these animal models, we could clarify the intrauterine transmission of HTLV-1, and have found that both genetic background and HTLV-1 infection are important in pathogenesis of HAM/TSP-like rats. We also discuss the preventive measures of HTLV-1 transmission by use of antisense oligonucleotides.

Abnormalities in late positive components of event-related potentials may reflect a genetic predisposition to schizophrenia.

Event-related potentials (ERPs) were recorded at the Cz region during syllable discrimination tasks in siblings of schizophrenic probands. ERPs in these siblings were compared to those of normal controls and schizophrenics. Siblings, similar to normal controls, displayed an increase in amplitudes of N100 according to the allocation of their attention between two different channels (ears), Siblings different from normal controls, however, failed to demonstrate an augmentation of late positive components upon detection of target stimuli in the attended channel. Mean amplitudes of late positive components elicited by target stimuli in the attended channel for siblings were nearly equal to those of unmedicated schizophrenics, with these values in siblings being significantly smaller as compared to those of normal controls. Based on these results, it was concluded that abnormalities of late positive components in siblings may reflect a genetic predisposition to schizophrenia.

The effects of analgesics on quickly-learned conditioned suppression in mice.

Mice were found to exhibit a marked suppression of motor activity when placed into the same experimental chamber in which they had previously received electrical shocks. This suppression was found to be a stable response when mice, 6-8 weeks of age, received shocks of frequencies of 1/10 or 1/30 Hz, and their motility was examined 24 hr after the delivery of the shock in the same experimental chamber. Control experiments indicated that the mice exhibited similar degrees of locomotor activity, compared to non-shocked mice, that were placed into a different experimental chamber than from the one in which they had previously received electrical shocks. Thus, the suppression of motor activity appeared to be a conditioned emotional response to the environment previously associated with delivery of shock. Morphine, codeine, and pethidine, but not pentazocine and aminopyrine, attenuated the conditioned suppression in a dose-related manner in the shocked mice. The effect of morphine was completely antagonized by pretreatment with naloxone or levallorphan. Thus, it is possible that the opiate-induced reduction of conditioned suppression is mediated by opiate receptor sites.

Effects of apomorphine and methamphetamine on a quickly-learned conditioned-suppression response in rats.

Rats exhibited a marked suppression of motor activity when placed in the same chamber where they had been given electric shocks. Administration of apomorphine-HCl (0.1, 0.5 and 1.0 mg/kg, i.p.) attenuated the conditioned suppression, dose-dependently, but did not facilitate motor activity of control (i.e. non-shocked) rats. Methamphetamine-HCl (0.1, 0.5 and 1 mg/kg, i.p.) increased motor activity of both the shocked and non-shocked rats, in a dose-related manner. Haloperidol (0.3 and 1 mg/kg, i.p.), but not chlorpromazine-HCl (5 mg/kg, i.p.), enhanced the conditioned-suppression response. Atropine-sulfate (5 and 10 mg/kg, i.p.), p-chlorophenylalanine (300 mg/kg, i.p.) and alpha-methyl-p-tyrosine (100 mg/kg, i.p.) were slightly effective in reducing the conditioned-suppression response. Apomorphine- and methamphetamine-induced reduction of the conditioned-suppression response was inhibited by pretreatment with haloperidol. When catecholamine-synthesizing processes in rats were inhibited by pretreatment with alpha-methyl-p-tyrosine, or alpha-adrenergic receptor sites were blocked by pretreatment with phenoxybenzamine, the effect of methamphetamine, but not that of apomorphine, was reduced. Therefore, enhancement of dopaminergic neurotransmission may be responsible for attenuation of the conditioned-suppression response.

The antagonistic effects of naloxone on hypermotility in mice induced by dynorphin-(1-13) using a multi-dimensional behavioral analysis.

The effects of intracerebral injection of dynorphin-(1-13) on spontaneous locomotor activity were investigated by using a newly devised multi-dimensional behavioural analyser based on a capacitance system. With this equipment, nine different measures of behavior exhibited by the mouse can be recorded. It was found that dynorphin-(1-13) (0.3 and 1 microgram) produced a significant increase in the linear locomotion recorded on the 1/1 and 1/2 counters. In addition, dynorphin-(1-13) (0.3 micrograms) significantly increased the grooming behavior recorded on the 1/16 counter. As the dynorphin-(1-13)-induced behavioural effects were reversed by pretreatment with relatively small doses of naloxone (0.5, 1 and 2 mg/kg), the dynorphin-induced increases in linear locomotion and grooming are probably mediated through opiate receptors in the brain.

DAMGO ([D-Ala2,NMePhe4,Gly-ol]enkephalin), but not DPLPE ([D-Pen2,L- Pen5]enkephalin), specifically inhibits methamphetamine-induced behavioral responses in the mouse.

The effects of intracerebroventricular (i.c.v.) injections of mu- and delta-selective opioid agonists on the methamphetamine-induced behavioral alterations in the mouse were determined by using multi-dimensional behavioral analyses. Methamphetamine (1.0 mg/kg) produced a marked increase in linear locomotion, circling, rearing and grooming behavior. Although the mu-selective opioid agonist [D-Ala2,NMePhe4,Gly-ol]enkephalin (DAMGO) (0.003 and 0.01 microgram) itself did not significantly affect different behavioral responses, DAMGO (0.003 and/or 0.01 microgram) antagonized the methamphetamine (1.0 mg/kg)-induced increase in behavioral responses such as linear locomotion, circling, rearing and grooming. Additionally, the effects of DAMGO (0.01 microgram) on the methamphetamine (1.0 mg/kg)-induced behavioral responses were fully reversed by pretreatment with the mu-selective alkylating agent beta-funaltrexamine (beta-FNA) (5.0 micrograms). In contrast, the delta-selective opioid agonist [D-Pen2,L-Pen5]enkephalin (DPLPE) (0.3 or 1.0 microgram) had no marked effects on the methamphetamine (1.0 mg/kg)-induced behavioral responses. These results suggest that the stimulation of mu but not delta opioid receptors plays an inhibitory role in the methamphetamine-induced behavioral responses.

Dynorphin A-(1-13) potently improves galanin-induced impairment of memory processes in mice.

The present study examined the effects of intracerebroventricular injection of dynorphin A-(1-13) on memory processes by using the passive avoidance task in mice. Galanin (0.3 microgram) significantly shortened the step-down latency when given 15 min before retention tests. Although dynorphin A-(1-13) (1 or 3 micrograms) did not prolong the step-down latency induced by weaker electroshocks, it inhibited the galanin (0.3 micrograms)-induced shortening of step-down latency. The effects of dynorphin A-(1-13) (3 micrograms) on the galanin-induced shortening of step-down latency were almost completely reversed by pretreatment with nor-binaltorphimine (4 micrograms), a kappa-selective opioid antagonist. These results strongly suggest that dynorphin A-(1-13) attenuates galanin-induced impairment of memory processes through the mediation of kappa-opioid receptors.

Effects of antidepressant drugs on a quickly-learned conditioned-suppression response in mice.

Mice experienced to electric shock, exhibited a marked suppression of motor activity when placed in the same cage 24 hr after administration of shocks. Acute administration of imipramine-HCl (10 mg/kg, i.p.), desipramine-HCl (5 and 10 mg/kg, i.p.) and amitriptyline-HCl (5 and 10 mg/kg, i.p.) caused marked reduction of the conditioned suppression of shocked mice, but reduced the motor activity of the non-shocked mice. Maprotiline, mianserin and dimetacrine did not cause reduction of the conditioned suppression. Nialamide (100 mg/kg, i.p.) and pargyline-HCl (100 and 200 mg/kg, i.p.) caused marked reduction of the conditioned suppression but did not increase the motor activity of the non-shocked mice, and tranylcypromine-HCl (10 and 20 mg/kg, i.p.) did not cause reduction of the conditioned suppression. Diphenhydramine-HCl (10 and 20 mg/kg, i.p.) reduced the conditioned suppression of shocked mice in a dose-related manner. Chronic administration of imipramine-HCl (1 and 5 mg/kg, i.p.) for 14 days significantly reduced the conditioned suppression but did not influence the motility rate of the non-shocked mice. Also, chronic administration of amitriptyline (1 mg/kg, i.p.), desipramine (5 mg/kg, i.p.) and dimetacrine (10 mg/kg, i.p.), for 10 days, significantly reduced the conditioned suppression, but did not influence the motility rate of the non-shocked mice. Chronic administration of maprotiline reduced the conditioned suppression. On the other hand, chronic administration of mianserin (5 mg/kg, i.p.) and diphenhydramine (10 mg/kg, i.p.) did not cause a reduction of the conditioned suppression.

The opioid antagonist, MR2266, specifically decreases saline intake in the mouse.

The effects of the opioid antagonist, Mr2266 [(-)-(1R,5R,9R)-5,9-diethyl-2-(3-furyl-methyl)-2'-hydroxy-6,7-benzomo rph an] on the intake of water and saline (0.9%) were investigated in the mouse, deprived of water for 24 hr. In an attempt to evaluate motor functions, the behavior after treatment with Mr2266 was also examined by using multi-dimensional behavioral analyses. Although smaller doses (1.0, 3.0 and 10.0 mg/kg) of Mr2266 failed to affect significantly the intake of water, a larger dose (30.0 mg/kg) elicited a significant attenuation in the intake of water. During a 30 min observation, Mr2266 (30.0 mg/kg) depressed markedly linear locomotion, while other behavioral responses, such as rearing and grooming, remained unchanged. In contrast, 1.0-30.0 mg/kg doses of the drug produced a significant reduction in the intake of saline. The drug Mr2266 had no significant effects on the latency to start drinking at any doses tested. These results suggest that Mr2266 specifically blocks the intake of saline of the mouse through the mediation of opioid systems.