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Long-term treatment with antiepileptic drugs (AEDs) is accompanied by reduced bone mass that is associated with an increased risk of bone fractures. Although phenytoin has been reported to adversely influence bone metabolism, little is known pertaining to more recent AEDs. The aim of this study was to evaluate the effects of gabapentin or levetiracetam on bone strength, bone mass, and bone turnover in rats. Male Sprague-Dawley rats were orally administered phenytoin (20 mg/kg), gabapentin (30 or 150 mg/kg), or levetiracetam (50 or 200 mg/kg) daily for 12 weeks. Bone histomorphometric analysis of the tibia was performed and femoral bone strength was evaluated using a three-point bending method. Bone mineral density (BMD) of the femur and tibia was measured using quantitative computed tomography. Administration of phenytoin significantly decreased bone strength and BMD, which was associated with enhanced bone resorption. In contrast, treatment with gabapentin (150 mg/kg) significantly decreased bone volume and increased trabecular separation, as shown by bone histomorphometric analysis. Moreover, the bone formation parameters, osteoid volume and mineralizing surface, decreased after gabapentin treatment, whereas the bone resorption parameters, osteoclast surface and number, increased. Levetiracetam treatment did not affect bone strength, bone mass, and bone turnover. Our data suggested that gabapentin induced the rarefaction of cancellous bone, which was associated with decreased bone formation and enhanced bone resorption, and may affect bone strength and BMD after chronic exposure. To prevent the risk of bone fractures, patients prescribed a long-term administration of gabapentin should be regularly monitored for changes in bone mass.

Long-term treatment with antiepileptic drugs (AED) is associated with an elevated risk of bone fracture due to decreased bone mineral density (BMD). Phenytoin has been shown to affect bone metabolism adversely, whereas newly developed AEDs have not been studied. This study evaluated the effects of topiramate and lamotrigine on bone metabolism in rats. Five-week-old male Sprague-Dawley rats were treated orally with phenytoin (20 mg/kg), topiramate (5 or 20 mg/kg), or lamotrigine (2 or 10 mg/kg) daily for 12 weeks. Phenytoin reduced bone strength, measured by maximum load to failure of the femoral mid-diaphysis, along with reduced femur total BMD. Serum tartrate-resistant acid phosphatase-5b levels significantly increased after phenytoin treatment, while serum osteocalcin levels decreased after topiramate 20 mg/kg treatment. Furthermore, osteoblast surface and bone mineralizing surface were significantly lowered by topiramate. Lamotrigine treatment did not affect bone strength, BMD, or bone turnover. We demonstrated that phenytoin treatment significantly increased bone resorption and lowered BMD and bone strength. Since lamotrigine did not affect bone metabolism, it can be concluded that lamotrigine is safety medicine for bone health. Topiramate was associated with decreased bone formation, which may affect bone strength and BMD with chronic use. Thus, patients taking topiramate should be monitored for changes in BMD to avoid risk of fracture.

The involvement of spinal glial cells in the nociceptive behaviors induced by 1600 pmol of histamine was determined in mice. Histamine injected intrathecally (i.t.) produced nociceptive behaviors, consisting of scratching, biting and licking. The nociceptive behaviors induced by histamine were significantly suppressed by i.t. pretreatment with the glial cell inhibitor DL-fluorocitric acid or minocycline. In Western blot analysis using lumber spinal cords, i.t. treatment with histamine increased the phosphorylation of the NR1 subunit of N-methyl-D-aspartate (NMDA) receptors. The increased phosphorylation of the NR1 subunit of NMDA receptors by histamine was abolished by i.t. pretreatment with DL-fluorocitric acid or minocycline. We have previously reported that the nociceptive behaviors induced by 1600 pmol of histamine were significantly suppressed by the i.t. co-administration of (5R,10 S)-(+)-5-methyl-10,11-dihydro-5 H-dibenzo[a,d]cycloheptene-5,10-imine (MK-801), an ion channel blocker of NMDA receptors, or agmatine, an antagonist for the polyamine recognition site on the NR1 subunit of NMDA receptors. In the present study, the increased phosphorylation of the NR1 subunit of NMDA receptors by histamine was also abolished by i.t. co-administration of agmatine or MK-801. The present results suggest that histamine at 1600 pmol elicits nociceptive behaviors by stimulating the polyamine recognition site on the NR1 subunit of NMDA receptors on spinal glial cells.

Zusammenfassung Mannliche Wistar-Ratten, welche mit thiaminfreiem Futter ernahrt wurden, zeigten Gewichtsverlust, Abfall der Herzfrequenz, Urinexkretionsstorungen und Symptome einer Polyneuritis. In dieser Studie wird berichtet, das Dauererektionen bei diesen Ratten vorkommen, das sich diese Erektionen wahrend des Versuchsablaufes fortsetzten, und das die Erektionen nach s.c. Injektion von Thiamin-HCl teilweise schwanden. Eine Ratte in der Thiamin-Mangelgruppe zeigte schon am 20. Versuchstag eine Dauer-erektion, wogegen dieser Effekt in dem Kontrollkollektiv nicht beobachtet wurde. Die Erektion war masig, der Penis ragte ca. 2 mm vor. Am 25. Tag wurde die Erektion deutlicher, das vorragende Penisstuck wurde ca. 5 mm lang. Wichtig ist, das die einmal ausgebildete Erektion wahrend der gesamten Versuchsdauer kontinuierlich anhielt. Am 30. Tag der Thiamin-Mangelfutterung zeigten 7 von 11 Ratten Dauererektionen, auserdem neurologische Erscheinungen wie Kreiselbewegungen und Nackensteife. Das Auftreten der Erektionen nahm standig zu, es betrug am 20. Tag 7%, am 25. Tag 30,7%, am 30. Tag 63,6% und am 33. Tag 100%. In der Thiamin-Mangelgruppe waren die Ratten mit Dauererektion unfahig zu urinieren, ihre Blasen waren vom gestauten Ham stark uberdehnt. Zusatzlich wurde in dieser Gruppe eine deutliche Einschrankung der Harnausscheidung beobachtet.

It has been reported that chlorpheniramine, a classical antihistamine, has antidepressant-like effects in animal models of depression. In this study, we examined the involvement of dopaminergic (dopamine D(1) and dopamine D(2) receptors), noradrenergic (alpha(1)- and beta-adrenoceptors) and serotonergic (5-HT(1A) and 5-HT(2) receptors) receptors in the antidepressant-like effect of chlorpheniramine in the mouse tail suspension test. We also investigated the involvement of these monoamine receptors in the antidepressant-like effect of imipramine for comparison with the mechanisms of the effect of chlorpheniramine. Both imipramine and chlorpheniramine significantly reduced the duration of immobility in the tail suspension test without affecting spontaneous locomotor activity in mice. The anti-immobility effect of imipramine (30 mg/kg, i.p.) was significantly antagonized by the selective dopamine D(1) receptor antagonist SCH23390 but not by the other receptor antagonists. In contrast, the anti-immobility effect of chlorpheniramine was significantly inhibited by SCH23390 and the selective alpha(1)-adrenoceptor antagonist prazosin, but not by the other receptor antagonists. In conclusion, these results suggest that chlorpheniramine exerts an antidepressant-like effect in the mouse tail suspension test that is mediated by at least the activation of dopamine D(1) receptors and alpha(1)-adrenoceptors. In addition, the antidepressant-like effect of chlorpheniramine may be induced by several mechanisms that are different from those involved in the antidepressant-like effect of imipramine.

Objectives We report a sensitive and selective method for the simultaneous determination of vitamin K1 and K2 analogs (VKs) with high-performance liquid chromatography in which a platinum catalyst reduction column and an electrochemical detector operated in the oxidation mode are incorporated into the detection system. We also applied this trace analysis method to the simultaneous determination of VKs in human serum to investigate the physiologic and pathophysiologic roles of VKs in the bone metabolism. Methods Our high-performance liquid chromatographic method with postcolumn catalyst reduction and electrochemical detection was applied for the simultaneous determination of VKs in human serum samples. After separation of VKs on a reversed-phase separation column by using a mixture of ethanol and methanol (1:1, v/v), containing 0.025 M of sodium perchlorate as the mobile phase, the VKs were reduced once in a platinum catalyst reduction column connected online and then monitored quantitatively by an electrochemical detector with a glassy carbon working electrode operated in the oxidation mode (+0.6 V versus Ag/AgCl). Results VKs were clearly separated from each other within 80 min. The detection limits at a signal-to-noise ratio of 3 were 2 to 10 pg for VKs. Quantitative recovery from serum was obtained in the range of 86% to 91% for VKs. The average coefficients of variation of within-day and between-day assays were 1.6% to 2.1% and 2.4% to 3.6%, respectively, for all VKs. Conclusions This method was sensitive and selective for detection of VKs and was satisfactory in the simultaneous determination of VKs in small volumes of human serum.

Objective: We investigated the effects of phenytoin, an antiepileptic drug, and vitamin K2 (menatetrenone) on bone mineral density and the changes in the levels of menaquinone derivatives (MK-1 ∼ MK-14) in the sera and femurs of growing male rats. Methods: Levels of menaquinone derivatives were measured with high-performance liquid chromatography with an electrochemical detector. Results: Bone mineral density values decreased significantly in all parts of the femoral bones measured (diaphysis and metaphysis) in the phenytoin-treated group. When the serum and bone levels of menatetrenone and MK-6 decreased due to phenytoin administration, we observed bone loss in rats. Conversely, when bone loss was prevented by the combined administration of phenytoin and menatetrenone, serum and bone levels of menatetrenone and MK-6 increased to the levels of vehicle-treated rats. Conclusions: Long-term phenytoin exposure may inhibit bone formation concomitantly with insufficient vitamin K, which, at least in part, contributes to bone loss in rats.

We investigated the effects of chronic administration of zonisamide, an antiepileptic agent, on bone metabolism in growing rats. Administration of zonisamide at a dose of 80 mg/kg per day, s.c. for 5 weeks significantly decreased bone mineral density (BMD) at the tibial metaphysis and the diaphysis. The percent rate of decrease in BMD at the tibial metaphysis and the tibial diaphysis was 9.2% and 5.0%, respectively. There was no significant difference between these groups in the growth of the rats. Treatment with zonisamide at a dose of 80 mg/kg increased serum pyridinoline level, a marker of bone resorption, while it does not affect the serum intact osteocalcin level, a marker of bone formation. Combined administration of alfacalcidol, an active vitamin D(3) metabolite, at a dose of 0.1 microg/kg per day with zonisamide prevented a decrease in BMD and showed an increase of serum pyridinoline levels. These results suggest that zonisamide may cause bone loss by accelerating bone resorption rather than inhibiting bone formation. Moreover, the bone loss induced by zonisamide could be prevented by combining zonisamide with alfacalcidol.

Intrathecal injection of histamine elicited a behavioral response consisting of scratching, biting and licking in conscious mice. Here, we have examined the involvement of substance P (SP) by using intrathecal injection of tachykinin neurokinin (NK)(1) receptor antagonists and SP antiserum. Histamine-induced behavioral response was evoked significantly 5-10 min after intrathecal injection and reached a maximum at 10-15 min. Dose-dependency of the induced response showed a bell-shaped pattern from 200 to 3200 pmol, and maximum effect was observed at 800-1000 pmol. The H(1) receptor antagonist, d-chlorpheniramine and pyrilamine but not the H(2) receptor antagonists, ranitidine and zolantidine, inhibited histamine-induced behavioral response. The NK(1) receptor antagonists, CP-99,994, RP-67580 and sendide, inhibited histamine-induced behavioral response in a dose-dependent manner. A significant antagonistic effect of [D-Phe(7), D-His(9)]SP (6-11), a selective antagonist for SP receptors, was observed against histamine-induced response. The NK(2) receptor antagonist, MEN-10376, had no effect on the response elicited by histamine. Pretreatment with SP antiserum resulted in a significant reduction of the response to histamine. No significant reduction of histamine-induced response was detected in mice pretreated with NK A antiserum. The present results suggest that elicitation of scratching, biting and licking behavior induced by intrathecal injection of histamine may be largely mediated by NK(1) receptors via H(1) receptors in the spinal cord.

The antinociceptive effects of pinacidil, an adenosine triphosphate (ATP)-sensitive K(+)i (K(ATP)) channel opener, were examined using the tail-flick test in non-diabetic and diabetic mice. Pinacidil i.c.v. produced dose-dependent antinociception in both non-diabetic and diabetic mice. There was no significant difference between the antinociceptive effect of i.c.v. pinacidil in non-diabetic mice and diabetic mice. The i.t. administration of pinacidil also produced dose-dependent antinociception in both non-diabetic and diabetic mice, however, the antinociceptive effect of i.t. pinacidil in diabetic mice was significantly greater than that in non-diabetic mice. The antinociceptive effect of i.c.v. or i.t. pinacidil was significantly antagonized by i.c.v. or i.t. glibenclamide, a K(ATP) channel blocker in both non-diabetic and diabetic mice. In non-diabetic mice, the antinociceptive effect of i.c.v. or i.t. administration of pinacidil was significantly antagonized by beta-funaltrexamine, a mu-opioid receptor antagonist, 7-benzylidenenaltrexone, a delta1-opioid receptor antagonist, naltriben, a delta2-opioid receptor antagonist, and nor-binaltorphimine, a kappa-opioid receptor antagonist. In diabetic mice, the antinociceptive effect of i.c.v. pinacidil was significantly reduced by 7-benzylidenenaltrexone, naltriben, and nor-binaltorphimine. However, beta-funaltrexamine had no effect on antinociception induced by i.c.v. pinacidil in diabetic mice. On the other hand, the antinociceptive effect of i.t. pinacidil was significantly antagonized by beta-funaltrexamine, 7-benzylidenenaltrexone, naltriben, and nor-binaltorphimine in diabetic mice. These results indicated that pinacidil produced antinociception through the release of opioid peptides acting at mu-, delta- and kappa-opioid receptors in surpraspinal and spinal cord of non-diabetic mice. On the other hand, in diabetic mice, the antinociception-induced by pinacidil was mediated through the release of opioid peptides acting at delta- and kappa-opioid receptors supraspinally, whereas pinacidil produced antinociception through the release of opioid peptides acting at mu-, delta-, and kappa-opioid receptors spinally.

In this study, we investigated 1) whether the administration of phenytoin induced bone loss; and 2) whether menatetrenone could prevent bone loss induced by phenytoin. For this purpose, we previously developed a procedure to measure the bone mineral density using a conventional X-ray absorptiometry method. A long-termed administration of phenytoin (20 mg/kg per day for 5 weeks) produced bone loss in the tibiae of growing rats. The values of bone mineral density (BMD) were significantly decreased in the tibial diaphysis and metaphysis in the phenytoin-treated group. In this period, we measured the serum level of vitamin K-dependent protein, osteocalcin, a marker of bone formation. The serum level of osteocalcin showed a decrease in the phenytoin-treated group compared with the vehicle-treated group. Combined administration of menatetrenone (30 mg/kg in diet per day) with phenytoin for 5 weeks prevented the reduction of BMD, and the level of osteocalcin was slightly increased. Thus, it is suggested that long-termed phenytoin exposure may inhibit bone formation concomitantly with insufficient vitamin K, which, at least in part, contributed to bone loss in rats. Finally, these findings implicated the therapeutic usefulness of menatetrenone on a moderate degree of bone abnormality such as drug-induced osteopenia.

Intrathecal (i.t.) injection of histamine elicited a significant hyperalgesic response as assayed by the tail-flick test. This hyperalgesic effect peaked at 15 min following i.t. administration of histamine (800 pmol) and returned to control level with 30 min. Hyperalgesia produced by histamine was inhibited dose-dependently by i.t. co-administration of the histamine H1 receptor antagonist, d-chlorpheniramine, but not the histamine H2 receptor antagonist, ranitidine. The tachykinin NK1 receptor antagonists, (+)-[(2S,3S)-3-(2-methoxy-benzyl-amino)-2-phenylpiperidine] (CP-99,994), and [Tyr6, d -Phe7, d -His9]substance P-(6–11) (sendide), inhibited histamine-induced hyperalgesic response in a dose-dependent manner. A significant antagonistic effect of [ d -Phe7, d -His9]substance P-(6–11), a selective antagonist for substance P receptors, was observed against histamine-induced hyperalgesic response. The tachykinin NK2 receptor antagonist, Asp-Tyr- d -Trp-Val- d -Trp- d -Trp-Lys-NH2 (MEN-10,376), had no effect on hyperalgesia elicited by histamine. The competitive N-methyl- d -aspartate (NMDA) receptor antagonists, and d -(−)-2-amino-5-phosphonovaleric acid ( d -APV), (±)-3-(2-carboxypiperazin-yl)propyl-1-phosphoric acid (CPP), the noncompetitive NMDA receptor antagonist dizocilpine (MK-801), and l -NG-nitro arginine methyl ester ( l -NAME), a nitric oxide (NO) synthase inhibitor, markedly inhibited histamine-induced hyperalgesic response. The present results suggest that hyperalgesic response induced by i.t. injection of histamine may be mediated by tachykinin NK1 receptors, but not NK2 receptors in the spinal cord. In addition, spinal NMDA receptor–NO system may also contribute to elicitation of hyperalgesia following i.t. injection of histamine.

We investigated whether inhaled pinacidil and moguisteine inhibit capsaicin-induced coughs in guinea pigs. Inhaled pinacidil (15 - 60 microg/ml), an ATP-sensitive K+ channel opener, and moguisteine (15 - 60 microg/ml) each dose-dependently inhibited the number of capsaicin-induced coughs. The antitussive effects of pinacidil and moguisteine were significantly antagonized by pretreatment with glibenclamide (10 mg/kg, i.p.), an ATP-sensitive K+ channel blocker. However, pretreatment with naloxone methiodide (10 mg/kg, s.c.) had no significant effect on the antitussive effects of either pinacidil or moguisteine. On the other hand, inhaled dihydrocodeine (15 - 60 microg/ml) also dose-dependently suppressed the number of capsaicin-induced coughs. The antitussive effect of inhaled dihydrocodeine was significantly antagonized by pretreatment with naloxone methiodide (10 mg/kg, s.c.), but not by glibenclamide (10 mg/kg, i.p.). These results indicate that inhaled pinacidil and moguisteine both attenuate capsaicin-induced coughs. Pinacidil and moguisteine may exert their antitussive effects through the activation of ATP-sensitive K+ channels in the tracheobronchial tract. Furthermore, it is possible that ATP-sensitive K+ channels may be involved in the antitussive effects of peripherally acting non-narcotic antitussive drugs.

We investigated the induction of metallothionein (MT) by cadmium (Cd) in the bone tissue of rats. To clarify the cell response to Cd in bone, the isoform-specific expression of MT mRNAs (MT-I and MT-II) was examined by reverse transcriptase-polymerase chain reaction (RT-PCR). Both MT-I and MT-II mRNA levels were increased within 3 h by Cd administration. MT (MT-I/MT-II) localization after single Cd injection were also confirmed by immunohistochemical studies. Notably, MT-positive cells were time-dependently increased, and the positive cells were mainly localized in osteocytes. The cell-specific induction of MT may be associated with Cd accumulation and Cd-induced bone injury in vivo. Furthermore, we also found that MT was consecutively expressed in some osteoclasts of control rats. This finding suggested a new role of osteoclasts in bone metabolism.

Studies were carried out to determine the effects and mechanism of action of phenytoin on the bone metabolism in male rats. Administration of phenytoin, 20 mg/kg/ day for 5 weeks, did not affect the growth curve. Biochemical data indicated that the serum osteocalcin, a marker of bone formation, was decreased significantly but there were no significant differences in the levels of serum calcium, pyridinoline, 25-hydroxyvitamin D3 (25OHD) and parathyroid hormone (PTH) in the phenytoin-treated group compared with the vehicle-treated group. The values of bone mineral density (BMD) were decreased in all regions of bones tested (mandibular head, tibial metaphysis, tibial diaphysis, femoral metaphysis, and femoral diaphysis) in the phenytoin-treated group. In histomorphometric analysis, phenytoin decreased trabecular bone volume and trabecular thickness, and increased osteoclast numbers per area of bone surface in the secondary trabecular bone of the tibia. Additionally, there was no significant difference in osteoid thickness. Combined administration of either alfacalcidol or calcitriol with phenytoin for 5 weeks prevented the reduction of BMD induced by phenytoin. From these findings, it is unlikely that toxic effects on the growth curve caused the decreased BMD induced by phenytoin. It is also evident that repeated administration of phenytoin may cause osteopenia which may be due to bone loss by inhibiting bone formation and/or by accelerating bone resorption rather than osteoid accumulation. The bone loss is not rachitic because of the lack of increase in osteoid thickness. Moreover, combined administration of alfacalcidol or calcitriol with phenytoin showed a preventative effect against bone loss. The bone loss induced by phenytoin in this study may be a convenient model for further research into the problem of drug-induced osteopenia.

The antinociceptive effect of vitamin K2 (menatetrenone) in mice was examined using tail-flick and formalin test. Menatetrenone at doses of 10, 50 and 100 mg/kg, i.p. produced a dose-dependent and significant inhibition of the tail-flick response in mice. Menatetrenone (50 and 100 mg/kg, i.p.) had no significant effect on the duration of the first phase of the formalin-induced flinching. However, menatetrenone (100 mg/kg, i.p.) significantly inhibited the second phase of the formalin-induced flinching. I.p. administration of menatetrenone (100 mg/kg) significantly reduced the duration of nociceptive responses induced by i.t. injection of bradykinin, but not of substance P, prostaglandin E2 or N-methyl-D-aspartate (NMDA). These present data suggest that i.p. pretreatment with menatetrenone produced dose-dependent antinociceptive effect in mice. This effect may be, at least in part, mediated by the inhibition of bradykinin dependent nociceptive transmission in the spinal cord.

We examined the effects of repeated exposure to forced walking stress for 6 h once a day for 0, 6 and 9 consecutive days on formalin-induced paw licking in mice. In each observation period, stress-induced antinociception (SIA) was observed only in the late phase (from 10 to 30 min), but not in the early phase (from 0 to 10 min) of formalin-induced paw licking in mice. Moreover, it was hard to develop tolerance even by daily exposure to stress for 6 days, although SIA for 9 days decreased compared with those for 0 and 6 days. Naloxone (10 mg/kg), an opioid-receptor antagonist, was effective in reducing the SIA induced by forced walking stress for 6 days and/or 9 days, but not for 0 days. Furthermore, the experiments with selective opioid-receptor antagonists, beta-funaltrexamine (mu) naltrindol (delta), or nor-binaltorphimine (kappa) demonstrated that SIA induced by forced walking stress for 9 successive days may be mediated through opioid delta- and kappa-receptors. Finally, although SIA seemed to be a unitary phenomenon, the present results strengthened the idea that SIA is induced by exposure to forced walking stress with characteristics dependent on the duration of exposure.

1. The purpose of this study was to compare the effect of NIK-247 on muscarinic receptor subtypes with that of tacrine (THA) in rats. 2. NIK-247 and tacrine dose dependently inhibited the binding of [3H]pirenzepine (M1), [3H]AF-DX 384 (M2), and [3H]4-DAMP (M3). The IC50 values for NIK-247 were 4.4 x 10(-6) M, 1.1 x 10(-5) M, and 1.5 x 10(-5) M, respectively, whereas those for tacrine were 5.8 x 10(-7) M, 2.0 x 10(-6) M, and 5.8 x 10(-6) M, respectively. 3. Gpp[NH]p, a GTP analogue, slightly shifted the curve of displacement of [3H]AF-DX. 384 binding for NIK-247 to the right. However, Gpp[NH]p did not shift the curve of displacement of [3H]pirenzepine and [3H]4-DAMP binding to the right. 4. NIK-247 moderately decreased the rate of beating in right atrial preparations, but did not decrease it below 50% of control level. 5. These findings indicate that NIK-247 is an M1 antagonist, M2 partial agonist, and M3 antagonist.

We recently demonstrated that some H-antagonists have cocaine or methamphetamine-like discriminative stimulus effects. In the present study, the effects of optical isomers of chlorpheniramine (D-, L- and DL-forms) on the discriminative stimulus effects of cocaine and methamphetamine were examined in rats trained to discriminate between cocaine (10.0 mg/kg) or methamphetamine (1.0 mg/kg) and saline, to determine whether these effects of H1-antagonists are mediated by the blockade of H-receptors. In generalization tests with optical isomers of chlorpheniramine, the D-, L- and DL-forms all completely generalized to the discriminative stimulus effects of cocaine, but did not generalize to those of methamphetamine. Dose-generalization by the optical isomers of chlorpheniramine to the discriminative stimulus effects of cocaine did not correlate with the H-antagonistic potency of these drugs. These results suggest that all of the optical isomers of chlorpheniramine have cocaine-like discriminative stimulus effects, but that these effects are not mediated by H1-receptor blockade. On the other hand, the H2-antagonist, zolantidine, generalized to the discriminative stimulus effects of methamphetamine, but not to those of cocaine, suggesting that zolantidine may have methamphetamine-like discriminative stimulus effects. In the present study, GBR12909 (dopamine uptake inhibitor) completely generalized to the discriminative stimulus effects of cocaine, but not to those of methamphetamine, whereas apomorphine (dopamine receptor agonist) generalized more potently to the discriminative stimulus effects of methamphetamine than to those of cocaine. These findings imply that although the dopaminergic system plays an important role in the discriminative stimulus effects of both cocaine and methamphetamine, there may be differences between their effects.

We studied the effect of therapeutic doses of theophylline on electrically-induced convulsions in developing mice. A theophylline dose as small as 3 mg/kg increased seizure susceptibility of 21-day-old mice, but not of 42-day-old mice. These findings were consistent with clinical reports that theophylline at the therapeutic blood concentrations occasionally induced convulsions in children. The age-dependent proconvulsant effect of theophylline was well inhibited by phenobarbital (PB), dose-dependently, but not by other well-established antiepileptic drugs (AEDs). PB may be a good choice of AED in patients with bronchial asthma and seizure disorders, if PB is indicative for their seizure types. The proconvulsant effect of theophylline in 21-day-old mice was counteracted by not only an adenosine A1 agonist, but also an NMDA antagonist and a histamine H3 antagonist. Several studies have established that the proconvulsant effect of theophylline intoxication is mainly due to the blockade of adenosine A1 receptors. The present findings suggested that the proconvulsant properties of therapeutic doses of theophylline in developing period were different from those of theophylline intoxication. Combination of therapeutic doses of theophylline and centrally-acting histamine H1 antagonists showed proconvulsant effects even in 42-day-old mice, suggesting that peripherally acting histamine H1 antagonists, such as astemizole, evastine and epinastine, were much safer than centrally acting histamine H1 antagonists for patients with both allergy and seizure history.

The effects of thioperamide, an H3 antagonist, and histamine H1 and H2 antagonists (s.c.) on morphine (s.c. or i.c.v.)- and U-50,488H (i.c.v.)-induced antinociception in male ddY mice were examined using the hot-plate (55 degrees C) test. Thioperamide significantly inhibited morphine-induced antinociception, but not U-50,488H-induced antinociception. The suppressive effect of thioperamide on morphine-induced antinociception was reversed by the H1 antagonist pyrilamine, but not by the H2 antagonist zolantidine. On the other hand, pyrilamine significantly potentiated the antinociception induced by morphine, but not that induced by U-50,488H. Zolantidine significantly inhibited morphine-induced antinociception in a dose-dependent manner, but not U-50,488H-induced antinociception. Both astemizole, an H1 antagonist, and ranitidine, an H2 antagonist, which are known to barely cross the blood brain barrier, did not affect morphine-induced antinociception. These results suggest that morphine-induced antinociception may be potentiated by activation of H2 receptors and suppressed by activation of H1 receptors in the brain. Furthermore, neuronal histamine release induced by thioperamide may suppress morphine-induced antinociception through H1 receptors.

The sonodynamically-induced in vitro and in vivo antitumor effects of mono-l-aspartyl chlorin e6 (NPe6) was investigated.Both in vitro and in vivo antitumor effects were tested in combination with ultrasound at 2 MHz.The rate of ultrasonically-induced damage on isolated sarcoma 180 cells in air-saturated suspension was enhanced two-fold with 80 μM NPe6. The co-administration of 25 mg/kg NPe6 followed by ultrasonic exposure at 2 MHz suppressed the growth of implanted colon 26 cell tumors at an intensity at which ultrasound alone showed only a slight antitumor effect.These in vitro and in vivo results suggest that NPe6 is a potential sensitizer for sonodynamic tumor treatment. The enhancement of cell damage by NPe6 was significantly inhibited by histidine, which may suggest reactive oxygen species plays a primary role in sonodynamically-induced antitumor effect.

The involvement of spinal glial cells in the nociceptive behaviors induced by 800 pmol of histamine was determined in mice. Histamine at 800 pmol injected intrathecally (i.t.) produced nociceptive behaviors, consisting of scratching, biting and licking. The nociceptive behaviors induced by histamine were significantly suppressed by i.t. co-administration with tachykinin NK(1) receptor antagonist CP99,994 or competitive antagonist for N-methyl-d-aspartate (NMDA) receptor d-(-)-2-amino-5-phosphonovaleric acid (d-APV). The i.t. pretreatment with the glial cell inhibitor dl-fluorocitric acid or minocycline failed to affect the nociceptive behaviors induced by histamine. However, in mice pretreated i.t. with dl-fluorocitric acid or minocycline, the nociceptive behaviors induced by histamine were significantly suppressed by i.t. co-administration with CP99,994 but not d-APV. In Western blot analysis using lumbar spinal cords, i.t. treatment with 800 pmol of histamine increased the phosphorylation of the NR1 subunit of NMDA receptors. The increased phosphorylation of the NR1 subunit of NMDA receptors by histamine was abolished by i.t. pretreatment with dl-fluorocitric acid or minocycline. The present results suggest that histamine at 800 pmol elicits nociceptive behaviors through activation of the neuronal NK(1) receptor and the NR1 subunit-containing NMDA receptors on glial cells in the spinal cord.

The purpose of this study was to investigate the developmental differences in seizure susceptibility in mice and the roles of the histaminergic neuron system in inhibition of convulsions in development. First, we studied developmental differences in electrically-induced seizures. Since the 14-day-old mice showed a different seizure pattern from that of older mice, we evaluated the seizure susceptibility of mice older than 21 days. The durations of all the convulsive phases were significantly increased in 21- and 30-day-old mice, compared with older mice. Second, pyrilamine (or mepyramine), ketotifen, and d-chlorpheniramine, centrally-acting H1-antagonists, increased the durations of all the convulsive phases in the 21- and 30-day-old mice, but did not increase duration in 42-day-old mice. Terfenadine and astemizole, H1-antagonists that hardly enter the brain, did not affect the durations of all the convulsive phases in 21-, 30- and 42-day-old mice. The proconvulsant effect of centrally-acting H1-antagonists in 21- and 30-day-old mice were considered to be mediated via the central H1-receptors. Thus, the histaminergic neuron system may have an important physiological role in inhibition of seizures in 21- and 30-day-old mice which have higher seizure susceptibility. This would compensate for the immaturity of the other protective neuron systems such as NMDA receptor complexes and GABA receptors. In conclusion, the present findings support the view that the central histaminergic system plays a role in inhibition of convulsions.

We studied the effects of inactivators of the central histaminergic neuron system, (R)-alpha-methylhistamine, a histamine H3 receptor agonist, and (S)-alpha-fluoromethylhistidine, a histamine synthesis inhibitor, on locomotor activity and brain histamine content of mast cell-deficient W/Wv mice using a recently developed high-performance liquid chromatography system coupled with a fluorometric detector. IP injection of (R)-alpha-methylhistamine (6-50 mg/kg) increased brain histamine content after 1 h but caused no significant change in locomotor activity. IP injection of (S)-alpha-fluoromethylhistidine decreased brain histamine content at doses of 6-50 mg/kg and locomotor activity at doses of 12.5-50 mg/kg. However, locomotor activity was decreased significantly (in Student's t-test) by sequential administrations of (S)-alpha-fluoromethylhistidine (6 mg/kg) and (R)-alpha-methylhistamine (12.5 or 25 mg/kg), but not by (S)-alpha-fluoromethylhistidine (6 mg/kg) and other doses of (R)-alpha-methylhistamine (6 or 50 mg/kg). These results support the hypothesis that the central histaminergic neuron system is involved in the control of spontaneous locomotion or alertness.

The antinociceptive effect of vitamin K2 (menatetrenone) in diabetic mice was examined using a tail-pressure test. Intraperitoneal injection of menatetrenone (10-100 mg/kg) produced a dose-dependent increase in the nociceptive threshold in diabetic mice. There was no significant difference between non-diabetic and diabetic mice in the menatetrenone-induced changes in the nociceptive threshold. The results suggest the therapeutical usefulness of menatetrenone for treating painful diabetic neuropathy and osteoporosis.

Studies were made on the role of μ1-receptors on electrically induced convulsions in mice. A relatively selective μ1-agonist, DAGO ([ d -Ala 2 -N- methyl-Phe 4 - Gly-ol]-enkephalin ) decreased the durations of tonic and clonic convulsions in C57BL/6 mice, but not in opioid μ-deficient CXBK mice, indicating that the activation of μ1-rreceptors had an anticonvulsive effect on these convulsions in mice. The selective μ1-antagonists naloxonazine and naltrexonazine promoted the clonic phase of electrically induced convulsion in C57BL/6 mice. Moreover, the duration of the clonic phase was longer in μ-deficient CXBK mice than in C57BL/6 mice. These data for CXBK mice confirmed, at least some proconvulsant effects of μ1-antagonists in C57BL/6 mice. Thus, blockade of μ1-receptors has a proconvulsant effect, and μ1-receptors have at least some protective role against electrically induced convulsions.

We examined the effects of (S)-alpha -fluoromethylhistidine (FMH), an inhibitor of histidine decarboxylase, and metoprine, an inhibitor of histamine N-methyltransferase, on the locomotor activity and the brain histamine content of ICR mice. The brain histamine content was decreased by FMH (12.5 or 50 mg/kg, i.p.) and increased by metoprine (4 mg/kg, i.p.). Under these conditions, the locomotor activity and the number of rearing were significantly decreased and increased by FMH and metoprine, respectively. The higher the brain histamine content, the greater the locomotor activity and vice versa. In a previous paper [Sakai et al., Life Sciences, 48, 2397-2404 (1991)], we showed that thioperamide, a histamine H3 antagonist, which enhances the release of histamine from histaminergic neurons, in doses of 12.5 and 25 mg/kg, i.p. increases the locomotor activity, whereas it decreases the brain histamine content. Taken together, these results support the hypothesis that central histaminergic neurons may be involved in the control of state of locomotion and rearing.

The purpose of this study was to further investigate the differences in the effects of optical isomers of chlorpheniramine (d-, l- and dl-forms) on the levels of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in the regional parts of rat brain. After the intravenous administration of each form of chlorpheniramine at a dose of 20 mg/kg, the DA and DOPAC levels were measured by HPLC system. Each form of chlorpheniramine is effective in reducing DOPAC, but not DA levels in the brain of rats. The degree of reducing DOPAC levels does not correlate with the antihistaminic potency of these drugs. Thus, the present results indicate that there is a lack of stereospecificity in reducing DOPAC levels, suggesting the lack of stereospecificity in inhibiting DA reuptake.

The rats maintained on a thiamine-deficient diet for 30 days showed hypothermia, and their histamine levels increased significantly in both the anterior and posterior hypothalamus. When these rats were administered, thiamine disulfide and/or provided with thiamine-added diet for a further 30 days, the rats recovered from hypothermia, and histamine levels were decreased to the normal level. Thus, it is probable that the increased histamine levels in the hypothalamus, especially those in its anterior region, are closely related to the hypothermia in thiamine-deficient rats.

In the present study, the effect of methamphetamine (MAP) on histamine (HA) release measured by in vivo brain microdialysis in the rat hypothalamus was investigated. Administration of MAP (3 mg/kg) significantly increase HA release from 40 to 160 min after the injection. This finding suggests that a moderate dose of MAP activates the hypothalamic HA neuron system, which may be related to effects of MAP on intrinsic biological rhythms.

We previously reported that streptozotocin-induced diabetic mice showed depressive-like behavior in the tail suspension test. It is well known that the central histaminergic system regulates many physiological functions including emotional behaviors. In this study, we examined the role of the central histaminergic system in the diabetes-induced depressive-like behavior in the mouse tail suspension test. The histamine contents in the hypothalamus were significantly higher in diabetic mice than in non-diabetic mice. The histamine H(1) receptor antagonist chlorpheniramine (1-10 mg/kg, s.c.) dose-dependently and significantly reduced the duration of immobility in both non-diabetic and diabetic mice. In contrast, the selective histamine H(1) receptor antagonists epinastine (0.03-0.3 microg/mouse, i.c.v.) and cetirizine (0.01-0.1 microg/mouse, i.c.v.) dose-dependently and significantly suppressed the duration of immobility in diabetic mice, but not in non-diabetic mice. Spontaneous locomotor activity was not affected by histamine H(1) receptor antagonists in either non-diabetic or diabetic mice. In addition, the number and affinity of histamine H(1) receptors in the frontal cortex were not affected by diabetes. In conclusion, we suggest that the altered neuronal system mediated by the activation of histamine H(1) receptors is involved, at least in part, in the depressive-like behavior seen in diabetic mice.

The second-generation histamine-H1-receptor antagonists, such as epinastine and cetirizine, are used as non-sedating antihistamines for treating allergic symptoms due to their poor ability to penetrate blood-brain barrier. Because it has been reported that the blood-brain barrier system is disturbed in diabetes, it is possible that second-generation histamine-H1-receptor antagonists may easily penetrate the blood-brain barrier and cause potent sedation in diabetics. In the present study, we investigated the effects of first-generation (diphenhydramine) and second-generation (epinastine and cetirizine) histamine-H1-receptor antagonists on the duration of pentobarbital-induced loss of the righting reflex (LORR) in non-diabetic and diabetic mice. Systemic treatment with diphenhydramine (3 – 30 mg/kg, s.c.), and intracerebroventricular treatment with epinastine (0.03 – 0.3 μg/mouse) and cetirizine (0.03 – 0.3 μg/mouse) dose-dependently and significantly increased the duration of pentobarbital-induced LORR in both nondiabetic and diabetic mice. Although systemic treatment with epinastine (3 – 30 mg/kg, s.c.) and cetirizine (3 – 30 mg/kg, s.c.) did not affect the duration of pentobarbital-induced LORR in nondiabetic mice, these treatments significantly prolonged it in diabetic mice. Our results suggest that the systemic administration of second-generation histamine-H1-receptor antagonists may produce a central nervous system depressant effect in diabetes. Keywords:: loss of righting reflex, histamine-H1-receptor antagonist, diabetes, pentobarbital, blood-brain barrier

The interactions between morphine and the histaminergic system are not yet fully clarified. More especially, the involvement of the histaminergic system in the discriminative stimulus effects of morphine has not been determined. Therefore, the effects of histamine-related compounds on the discriminative stimulus effects of morphine were examined in rats. Combination tests using histamine-related compounds with morphine were initiated in rats trained to discriminate between 3.0 mg/kg morphine and saline. Zolantidine (central histamine H2-receptor antagonist), but not pyrilamine (central histamine H1-receptor antagonist) or ranitidine (peripheral histamine H2-receptor antagonist), significantly attenuated the discriminative stimulus effects of morphine. The histamine precursor l -histidine significantly potentiated the discriminative stimulus effects of morphine. These results suggest that the discriminative stimulus effects of morphine are, at least in part, mediated through the central activation of histamine H2-receptors in rats.

We investigated the involvement of adenosine receptors on forced walking stress-induced analgesia using a formalin-induced paw-licking test in male mice. Exposure to forced walking stress for 6 h showed stress-induced analgesia in the second phase (10-30 min), but not in the first phase (0-10 min). In the second phase, forced walking stress-induced analgesia was blocked by theophylline, a nonselective adenosine-receptor antagonist and DPCPX, an adenosine A1-receptor antagonist, but not ZM 241385, an adenosine A2A-receptor antagonist. These findings suggest that adenosine A1 receptors are involved in the analgesic mechanism activated by the forced walking stress.

Background: Tartrate-resistant acid phosphatase (TRAP) is known as a marker of bone resorption. The purpose of this study was the development of a sensitive and specific immunoassay for TRAP. Methods: We have developed two types of immunoassays, enzyme-linked immunosorbent assay (ELISA) and immunoselective enzyme immunoassay (ISEA) using monoclonal antibodies to recombinant TRAP, for determination of TRAP in human serum. To evaluate assay performance, recovery and dilution tests were performed. Further, we determined serum TRAP levels of patients with secondary hyperparathyroidism at different pH conditions. Results: The detected ranges of ELISA and ISEA were between 0.08 and 5 μg/l and between 0.063 and 4 U/l. Different concentrations of TRAP added were recovered on average at 98.0% in ELISA and 102.9% in ISEA. In the serial dilution test, serum TRAP levels were on average at 101.6% and 109.6% of the expected values in ELISA and ISEA, respectively. The serum TRAP levels of patients with secondary hyperparathyroidism were significantly higher than those of normal controls in ELISA and ISEA. Similar TRAP levels were obtained in the conditions at pH 5.5 and 6.1 in ISEA. Conclusion: The present findings suggest that our assay methods are applicable for clinical tests, and strengthen the idea that serum TRAP is useful as a marker for bone resorption.

A study of the different volume and infusion rates of a new maintenance fluid, Veen 3G, on the general conditions of rats was investigated during the 14 days after infusion. In Experiment I, 100 ml/kg and 200 ml/kg of Veen 3G were infused at a rate of 300 ml/kg/h in male and female rats. Results were compared with those for Gurunon Ringer solution (GRS) in male and female rats. We observed only transient polyuria in animals administered by each dose of Veen 3G and GRS for 0-15 min after infusion. Necropsy was not observed in any of the animals tested 14 days after infusion. In Experiment II, 200 ml/kg of Veen 3G was infused at rates of 200, 400, 800 and 1600 ml/kg/h in male rats. At 800 and 1600 ml/kg/h, irregular respiration and decrease in movement were observed concomitantly with polyuria. Three out of 4 rats died immediately after the infusion of Veen 3G at a rate of 1600 ml/kg/h, and one rat was still alive 14 days after the infusion. In this experiment, 200 ml/kg Veen 3G was safe when we infused at a rate of less than 400 ml/kg/h in male rats. Since this rate is about 27-80 times higher than that used clinically in maintenance treatment, Veen 3G is suggested to be safe, with the exception of polyuria, in clinical situations at the standard infusion rate (5-15 ml/kg/h).

The role of the histaminergic system in the discriminative stimulus effects of cocaine and methamphetamine was examined in rats trained to discriminate between saline and cocaine (10 mg/kg) or methamphetamine (1.0 mg/kg). L-histidine (400 mg/kg), a precursor of histamine, significantly enhanced the discriminative stimulus effects of cocaine and methamphetamine. Previous studies have revealed the existence of several histamine receptor types, H1-, H2-, and H3-receptors. These enhancing effects of L-histidine on the discriminative stimulus effects of cocaine and methamphetamine were attenuated by 5.0 mg/kg of pyrilamine (an H1-receptor antagonist), but not by 1.0 mg/kg of zolantidine (an H2-receptor antagonist), suggesting that these enhancing effects of L-histidine were mediated through the activation of H1-receptors. Thioperamide (7.5 mg/kg), an H3-receptor antagonist, also significantly enhanced the discriminative stimulus effects of cocaine and methamphetamine. However, neither pyrilamine nor zolantidine affected the enhancing effects of thioperamide, unlike the results attained with L-histidine. Therefore our findings suggest that the histaminergic system may modify the discriminative stimulus effects of cocaine and methamphetamine mediated through H1- and H3-receptors.

Little information is available about the interaction between dihydrocodeine and second-generation antihistamine drugs such as cetirizine and ebastine, with particular reference to the rewarding effect of dihydrocodeine.The effects of second generation histamine H(1) antagonists, such as cetirizine and ebastine on the antitussive and rewarding effect of dihydrocodeine were examined in mice.Mice were exposed to a nebulized solution of capsaicin (30 micromol/l) under conscious and identical conditions, using a body plethysmograph. The coughs produced during a 3-min exposure period were counted. Effects of H(1) antagonists on the reinforcing effect of dihydrocodeine were assessed by using the conditioned place preference procedure in mice.The antitussive effect of dihydrocodeine was enhanced by the simultaneous administration of either cetirizine or ebastine. There was no statistical difference between the ED(50) of dihydrocodeine in combination with ebastine and that of dihydrocodeine in combination with cetirizine. Concurrent dosing of dihydrocodeine and ebastine produced a significant place preference. This behavioral potentiation was antagonized by SCH23390, a dopamine D(1) antagonist. Moreover, ebastine enhanced the central dopamine turnover ratio, but cetirizine could not, in this study.Taken together, the potentiation of place preference of dihydrocodeine with ebastine may be due, at least in part, to stimulation of the central dopaminergic system via D(1) receptors. However, combination of dihydrocodeine with cetirizine does not potentiate place preference at all, nor does it potentiate the central dopaminergic system. Thus, it is likely that cetirizine may be a useful constituent in opioid-containing, antitussive preparations that would not potentiate the development of psychological dependence.

We examined the effect of prazosin, a selective alpha1-adrenergic receptor antagonist, on the enhanced spontaneous locomotor activity in streptozotocin-induced diabetic mice. Spontaneous locomotor activity in diabetic mice was significantly greater than that in nondiabetic mice. Pretreatment with either intracerebroventricular (5, 10 nmol) or intraperitoneal (0.5, 1.0 mg/kg) injection of prazosin dose-dependently reduced the spontaneous locomotor activity in diabetic mice, but not in nondiabetic mice. Furthermore, the enhanced dopamine turnover ratio in the limbic forebrain in diabetic mice was reduced to the same level as that in nondiabetic mice after the administration of prazosin. Thus, these results suggest that alpha1-adrenergic receptors might play an important role in the enhanced spontaneous locomotor activity in diabetic mice. Furthermore, alpha1-adrenoceptor antagonism might have an inhibitory effect on presynaptic dopaminergic neurotransmission in the limbic forebrain in diabetic mice.

The purpose of this study was to evaluate the relationship between the in vivo toxicity and plasma concentration of theophylline. Theophylline was administered intravenously in single doses ( 50, 100, 150 and 200 mg kg −1 once a day) or repeated doses (12.5, 25 and 90 mg kg −1 /day for 28 days) in rats. Plasma concentrations of theophylline increased dose-dependently in both single and repeated doses, and there were no differences due to effects of 28-times repeated administration. Neither single dose at 50 mg kg −1 nor repeated dose at 12.5 mg kg −1 /day injections of theophylline showed toxic signs, in which plasma concentrations of theophylline were less than 110 and 22.5 μ g ml −1 , respectively. Theophylline induced myocardial fibrosis in 25 mg kg −1 /day and more treated groups: in which plasma concentrations of theophylline were more than 50 μ g ml −1 . At doses of 100 mg kg −1 (single) and 90 mg kg −1 /day (repeated), theophylline caused tachypnea and excitement of movement. Each theophylline concentration in plasma was more than 194 μ g ml −1 in single 100 mg kg −1 and 162 μ g ml −1 in repeated 90 mg kg −1 /day injections, respectively. Death was observed at a dose of 200 mg kg −1 , in which the plasma concentration of theophylline was more than 264 μ g ml −1 . Moreover, the recovery period from signs of toxic poisoning to normality in the 200 mg kg −1 treated group was greater than that in the 150 mg kg −1 and less treated groups. The results indicated that the in vivo toxicity of theophylline is highly dependent on plasma concentrations in rats which received single and also repeated doses of theophylline.

We investigated the induction of metallothionein (MT) by cadmium (Cd) in the dental pulp of rat incisors. Time-course studies of MT mRNA expression after single Cd injection were observed by Northern-blot analysis. The isoform-specific expressions of MT mRNAs (MT-I, MT-II and MT-III) were observed using the reverse transcriptase-polymerase chain reaction (RT-PCR) method. Both MT-I and MT-II mRNA levels increased within 3 h, peaked at 3 h and then decreased. These findings demonstrated that MT-I and MT-II mRNA were rapidly induced by Cd in dental pulp. MT-III mRNA was constitutively expressed in rat dental pulp, but the expression level did not change by Cd treatment. The localization of MT protein in Cd-treated rat dental pulp was determined by immunohistochemical staining using anti-MT antibody against MT-I and MT-II. MT protein was localized in the specific cell type of odontoblasts (secretory odontoblasts and resting odontoblasts). In conclusion, it is likely that stained MT in the immunohistochemical study should be MT-I and/or MT-II. Furthermore, MT-I and/or MT-II in Cd-treated rat dental pulp was localized in odontoblasts, in which accumulation of Cd were reported. The cell-specific synthesis of MT may be associated with its metal storage and detoxification role in dental tissues.

Metallothioneins (MTs) have a low molecular weight and have been considered to be important metal-binding proteins in defense from cadmium (Cd) toxicity in animals. These proteins are known to be induced by the injection of heavy metals such as Cd. Previously, we developed the measurement of the MT mRNA expression by the RT-PCR method. In this study, to clarify the relation between Cd-induced MT-I mRNA expression and female sex hormones in liver, we investigated the influences of the ovariectomy and female sex hormones on hepatic MT-I mRNA expression after Cd injection, and also investigated the effects of aging on hepatic MT-I mRNA expression in mice after Cd injection. We analysed the MT-I mRNA expression by the RT-PCR method. Cd-induced MT-I mRNA expression in ovariectomized mice was more than that in sham-operated mice (9 weeks old). Both 17 β -estradiol and progesterone reduced the Cd-induced MT-I mRNA expression in ovariectomized mice (9 weeks old). Moreover, the MT-I mRNA expression in male mice was more than that of females (9 weeks old). However, the sex difference in the gene expression was not found in younger (4 weeks old) or older (46 weeks old) mice. These results suggest that the expression of hepatic MT-I mRNA after Cd injection is influenced by female sex hormones.

Metallothioneins (MTs) are low molecular weight proteins that have been considered to be important metal-binding proteins in the defense against cadmium (Cd) toxicity in animals. These proteins are known to be induced by the injection of heavy metals such as Cd. Previously, we developed the RT-PCR method to measure the level of MT mRNA expression. In this study, we analyzed the time course and dose response of Cd-induced MT-I mRNA expression in the male and female mouse liver. By this method, we measured hepatic MT-I mRNA expression which reached the highest peak 6 h after subcutaneous injection of 1.1 mg/kg Cd in 9-week old male and female mice compared with those of corresponding controls (0 h). There was no statistical difference in the hepatic MT-I mRNA expression between male and female mice 6 h after the injection of this dosage. However, it is notable that the MT-I mRNA expression in male mice was much higher than that in females 6 h after injection of 0.5 mg/kg Cd. Thus, the induction of hepatic MT-I mRNA expression is dependent on the sex of the mouse, the dosage and the time course of Cd.

This study investigated the teratogenic and fetal toxicity of i.v. theophylline and its relationship to maternal plasma levels in pregnant rabbits. From days 6-18 of gestation, each dose of theophylline (15, 30 and 60 mg/kg/day at a rate of 0.5 ml/kg/min) was administered i.v. to pregnant rabbits using an automatic infusion pump. Theophylline showed reversible toxicity: accelerated respiration, sluggish startle reactions, dilation of the auricular vessels and polyuria were observed in dams treated with 60 mg/kg/day but not in animals given 15 or 30 mg/kg/day. Fetuses from the dam group treated with 60 mg/kg/day exhibited teratogenic toxicity such as cleft palate and skeletal variation of the 13th rib. Fetal toxicity was also observed including abortion, increased number of late deaths and decreased body weight appearing on day 29 of gestation. No toxicity was observed in fetuses from the dam group treated with 15 or 30 mg/kg/day. However, in the 30 and 60 mg/kg/day theophylline-treated groups, maternal plasma concentrations (Cmax) during the treatment period were approximately 56 and 106 micrograms/ml, respectively. It is therefore suggested that the risk of teratogenic and fetal toxicity caused by theophylline is dependent on its dosage. In conclusion, caution should be taken when administering theophylline or aminophylline to pregnant individuals at doses that could result in high neonate peak blood levels.

We investigated behavioral changes in mice in a light/dark test after forced swimming stress in a streaming pool. Locomotion and rearing behavior in mice in a light/dark test was suppressed immediately after the swimming and gradually regained. Shuttle crossing between light and dark zones was also suppressed but time spent in a light zone was not. The suppression of locomotor activity by the swimming was not observed 20 min after it. Naloxone (10 mg/kg, i.p.) suppressed recovery of locomotor activity after the swimming while it did not affect a light/dark test in nonswimming mice. These results suggest that this model is useful for the study of behavioral and psychological changes after exercise, and that opioidergic systems may be involved in regaining spontaneous locomotor activity.

1. The purpose of this study was to examine whether NIK-247 can, by itself, induce long-lasting changes in synaptic efficacy in the hippocampus. Population spikes evoked by electrical stimulation of the stratum radiatum were recorded in the pyramidal cell layer of the CA1 region of the isolated hippocampus. 2. NIK-247 at 1×10−7–1×10−5 M dose dependently increased the amplitude of these spikes. The increase in population spikes by NIK-247 outlasted, for 2 hr, its presence. In addition, the increase in population spikes recovered to 2 hr after washout of NIK-247. Therefore, it was concluded that NIK-247 induced long-term potentiation (LTP) by itself. However, tacrine and physostigmine at 1×10−7–1×10−5 M did not increase the amplitude of population spikes and did not induce LTP by themselves. 3. The increase in amplitude of population spikes induced by NIK-247 was completely blocked sensitively by atropine IC 50 =4.3 × 10 −8 M but insensitively by pirenzepine IC 50 =9.1 × 10 −7 M . Carbachol also increased the amplitude of population spikes in the presence of pirenzepine. 4. These findings indicate that the LTP induced by NIK-247 is due to its M2 muscarinic agonistic effect in the CA1 region of the rat hippocampus. It is expected that NIK-247 may be useful for the treatment of Alzheimer disease.

Effects of FUB 181 [3-(4-chlorophenyl)propyl-3-(1H-imidazol-4-yl)propyl ether], a novel histamine H3-receptor antagonist, on a scopolamine-induced learning deficit in the elevated plus-maze test were studied in mice. FUB 181 alone (2.5 and 5 mg/kg, i.p.) ameliorated the scopolamine-induced learning deficit in mice. This effect was antagonized by BP 2.94 (10 mg/kg, i.p.), a prodrug of (R)-alpha-methylhistamine (histamine H3-receptor agonist), and by ketotifen (4 mg/kg, i.p.), a histamine H1-receptor antagonist, both penetrating the blood-brain barrier. However, the ameliorating effect of FUB 181 (2.5 mg/kg) was not antagonized by either terfenadine (10 mg/kg, i.p.), a histamine H1-receptor antagonist with poor penetration of the blood-brain barrier, or zolantidine (20 mg/kg, i.p.), a centrally effective histamine H2-receptor antagonist. In a biochemical study, FUB 181 had no significant effect on either acetylcholine or choline level in mice brain at the doses tested. These findings suggest that FUB 181 increases the release of histamine by blocking presynaptic histamine H3 autoreceptors, and that released histamine in turn activates postsynaptic H1 and H2 receptors, predominantly histamine H1 receptors, and in this fashion improves learning and memory in mice. Our findings also suggest that the histaminergic system may play an important role in learning and memory, and that FUB 181 may be a clinical candidate for the therapy of dementia.

We previously demonstrated that combination of opioids, pentazocine and dihydrocodeine, with the histamine H1-receptor antagonists tripelennamine and chlorpheniramine could enhance their rewarding effects in rats. In the present study, the effects of combined treatment with opioids and H1-antagonists on discriminative stimulus effects were examined in rats trained to discriminate between cocaine (10 mg/kg) or morphine (3.0 mg/kg) and saline, since it is believed that discriminative stimulus effects of abused drugs are related to their rewarding effects. Tripelennamine and chlorpheniramine, but not pentazocine or dihydrocodeine, generalized to the discriminative stimulus effects of cocaine. Pentazocine (3.0 mg/kg) and dihydrocodeine (5.6 mg/kg) significantly potentiated the cocaine-like discriminative stimulus effects of low doses of tripelennamine and chlorpheniramine, respectively. On the other hand, pentazocine and dihydrocodeine, but not tripelennamine or chlorpheniramine, generalized to the discriminative stimulus effects of morphine. Neither 1.0 or 3.0 mg/kg tripelennamine nor chlorpheniramine affected the morphine-like discriminative stimulus effects of pentazocine and dihydrocodeine, respectively. These results suggest that the potentiation of the cocaine-like discriminative stimulus effects of H1-antagonists by opioids may be involved in the enhanced rewarding effects of combinations of opioids and H1-antagonists.

In this study, effects of histamine (HA) agents on methamphetamine (METH)-induced stereotyped behavior and behavioral sensitization were examined in rats. Pretreatment with a precursor of HA, L-histidine (750mg/kg), significantly inhibited the METH (3 mg/kg)-induced stereotyped behavior, whereas pretreatment with an inhibitor of HA synthesis, alpha-fluoromethylhistidine (FMH) (100 mg/kg), an H1 antagonist pyrilamine (5 mg/kg) or an H2 antagonist zolantidine (5 mg/kg) enhanced it. The inhibitory effect of L-histidine on METH-induced stereotyped behavior was significantly blocked by coadministration of pyrilamine and zolantidine, indicating that the effect is mediated through H1 and H2 receptors. Moreover, chronic treatment with METH (3 mg/kg) significantly enhanced stereotyped behavior at the rechallenge with METH (1 mg/kg). Chronic treatment with L-histidine (750 mg/kg) plus METH inhibited the METH-induced argumentation of stereotyped behavior, while that with FMH (100 mg/kg), pyrilamine (5 mg/kg) or zolantidine (5 mg/kg) potentiated it. These findings suggest that the HA neuron system has an inhibitory role in METH-induced stereotyped behavior and behavioral sensitization.