Overexpression of E2F-5 correlates with a pathological basal phenotype and a worse clinical outcome.

The purpose of the present study is to identify genes that contribute to cell proliferation or differentiation of breast cancers independent of signalling through the oestrogen receptor (ER) or human epidermal growth factor receptor 2 (HER2). An oligonucleotide microarray assayed 40 tumour samples from ER(+)/HER2(-), ER(+)/HER2(+), ER(-)/HER2(+), and ER(-)/HER2(-) breast cancer tissues. Quantitative reverse transcriptase PCR detected overexpression of a cell cycle-related transcription factor, E2F-5, in ER-negative breast cancers, and fluorescence in situ hybridisation detected gene amplification of E2F-5 in 5 out of 57 (8.8%) breast cancer samples. No point mutations were found in the DNA-binding or DNA-dimerisation domain of E2F-5. Immunohistochemically, E2F-5-positive cancers correlated with a higher Ki-67 labelling index (59.5%, P=0.001) and higher histological grades (P=0.049). E2F-5-positive cancers were found more frequently in ER(-)/progesterone receptor (PgR)(-)/HER2(-) cancer samples (51.9%, P=0.0049) and in breast cancer samples exhibiting a basal phenotype (56.0%, P=0.0012). Disease-free survival in node-negative patients with E2F-5-positive cancers was shorter than for patients with E2F-5-negative cancers. In conclusion, we identify, for the first time, a population of breast cancer cells that overexpress the cell cycle-related transcription factor, E2F-5. This E2F-5-positive breast cancer subtype was associated with an ER(-)/PgR(-)/HER2(-) status, a basal phenotype, and a worse clinical outcome.

Pharmacokinetics of zonisamide and drug interaction with phenobarbital in dogs.

The purposes of the present study were to elucidate the pharmacokinetics of zonisamide, determine the presence of a drug interaction with phenobarbital, and evaluate how long any interaction lasted after discontinuation of phenobarbital in dogs. Five dogs received zonisamide (5 mg/kg, p.o. and i.v.) before and during repeated oral administration of phenobarbital (5 mg/kg, bid, for 30-35 days). Zonisamide (5 mg/kg, p.o.) was also administered 8, 10, and 12 weeks after discontinuation of phenobarbital. Blood was sampled until 24 h after each zonisamide administration and serum concentrations of zonisamide were determined. Repeated phenobarbital decreased the maximum serum concentration, area under the serum concentration vs. time curve, apparent elimination half-life, and bioavailability of zonisamide. Total clearance increased. Time to maximum serum concentration and volume distribution were not changed. The maximum serum concentration and area under the serum concentration vs. time curve of zonisamide continued to be low until 10 weeks after the discontinuation of phenobarbital. They were restored to the same serum concentration as before phenobarbital administration 12 weeks after the discontinuation of phenobarbital. These data suggested that repeated administration of a clinical dose of phenobarbital enhanced the clearance of zonisamide and the enhanced clearance lasted at least 10 weeks after the discontinuation of phenobarbital. Caution may be necessary when zonisamide is given with phenobarbital and when antiepileptic therapy is changed from phenobarbital to zonisamide.

Delta 9-tetrahydrocannabinol-induced catalepsy-like immobilization is mediated by decreased 5-HT neurotransmission in the nucleus accumbens due to the action of glutamate-containing neurons.

Delta(9)-tetrahydrocannabinol (THC) has been reported to induce catalepsy-like immobilization, but the mechanism underlying this effect remains unclear. In the present study, in order to fully understand the neural circuits involved, we determined the brain sites involved in the immobilization effect in rats. THC dose-dependently induced catalepsy-like immobilization. THC-induced catalepsy-like immobilization is mechanistically different from that induced by haloperidol (HPD), because unlike HPD-induced catalepsy, animals with THC-induced catalepsy became normal again following sound and air-puff stimuli. THC-induced catalepsy was reversed by SR141716, a selective cannabinoid CB(1) receptor antagonist. Moreover, THC-induced catalepsy was abolished by lesions in the nucleus accumbens (NAc) and central amygdala (ACE) regions. On the other hand, HPD-induced catalepsy was suppressed by lesions in the caudate putamen (CP), substantia nigra (SN), globus pallidus (GP), ACE and lateral hypothalamus (LH) regions. Bilateral microinjection of THC into the NAc region induced catalepsy-like immobilization. This THC-induced catalepsy was inhibited by serotonergic drugs such as 5-hydroxy-L-tryptophan (5-HTP), a 5-HT precursor, and 5-methoxy-N,N-dimethyltryptamine (5-MeODMT), a 5-HT receptor agonist, as well as by anti-glutamatergic drugs such as MK-801 and amantadine, an N-methyl-d-aspartate (NMDA) receptor antagonist. THC significantly decreased 5-HT and glutamate release in the NAc, as shown by in vivo microdialysis. SR141716 reversed and MK-801 inhibited this decrease in 5-HT and glutamate release. These findings suggest that the THC-induced catalepsy is mechanistically different from HPD-induced catalepsy and that the catalepsy-like immobilization induced by THC is mediated by decreased 5-HT neurotransmission in the nucleus accumbens due to the action of glutamate-containing neurons.

MRI appearance of primary giant ovarian leiomyoma in a hysterectomised woman.

Primary leiomyoma is a rare, benign tumour of the ovary. We describe the MRI features of an ovarian leiomyoma identified in a 51-year-old woman after hysterectomy. The tumour appeared as a well-circumscribed low signal intensity mass on T(1) weighted imaging, with mixed signal intensity on T2 weighted imaging. Areas of high signal intensity on T2 weighted imaging corresponded to degeneration of leiomyoma. Dynamic contrast-enhanced imaging revealed early enhancement of the lesion. MRI appearance was thus similar to that of uterine leiomyoma. This case suggests the potential usefulness of dynamic contrast-enhanced MRI for the diagnosis of ovarian leiomyoma.

Ninjin-yoei-to (Ren-Shen-Yang-Rong-Tang) and Polygalae radix improves scopolamine-induced impairment of passive avoidance response in mice.

We investigated the effect of Ninjin-yoei-to (Ren-Shen-Yang-Rong-Tang), a Japanese herbal medicine, and found that 1000 mg/kg p.o. improved the scopolamine-induced impairment of passive avoidance response in mice. Further, the same dose of Ninjin-yoei-to enhanced oxotremorine-induced tremors in mice. The water extract of Polygalae radix, one of the constituent herbs of Ninjin-yoei-to, at a dose of 100 mg/kg significantly improved the scopolamine-induced impairment of passive avoidance response and enhanced oxotremorine-induced tremors in mice. Moreover, the enhancement of oxotremorine-induced tremors by Ninjin-yoei-to (1000 mg/kg) and Polygalae radix (100 mg/kg) was completely antagonized by pretreatment of scopolamine hydrobromide (0.5 mg/kg). These results suggest that Ninjin-yoei-to may improve the scopolamine-induced impairment of passive avoidance response by enhancing the cholinergic system and that Polygalae radix may be involved in the action of Ninjin-yoei-to.

Post-ischemic administration [correction of administeration] but not pre-ischemic administration [correction of administeration] of NG-nitro-L-arginine prevents spatial memory impairments and apoptosis by an inhibition of a delayed increase in NOx- in the hippocampus following repeated cerebral ischemia.

In the present study, we investigated the effects of N(G)-nitro-L-arginine (L-NAME), an inhibitor of nitric oxide synthase, on repeated cerebral ischemia-induced impairment of spatial memory of the 8-arm radial maze in rats. Repeated ischemia (10 min ischemia x 2 times with 1 h interval) impaired the spatial memory in the 8-arm radial maze test and produced apoptosis in the hippocampus 7 days after final occlusion, and gradually increased the NO(x)(-) levels approximately 30-180 min after the second reperfusion. Post-ischemic administration of L-NAME at a dose of 50 mg/kg, i.p. 30 min following the second occlusion, significantly attenuated the repeated ischemia-induced impairment of spatial memory in the 8-arm radial maze test and suppressed apoptosis in the hippocampus, and also significantly suppressed a delayed increase in the NO(x)(-) levels induced by repeated ischemia. However, pre-ischemic administration of L-NAME at a dose of 50 mg/kg, i.p. 30 min before the first occlusion, caused about 90% mortality (the mortality rate of vehicle-treated group was 10%). These results suggest that the delayed generation of NO(x)(-) may cause spatial memory impairment and induction of apoptosis in the hippocampus in rats subjected to repeated ischemia.

Ameliorative effect of NC-1900, a new AVP4-9 analog, through vasopressin V1A receptor on scopolamine-induced impairments of spatial memory in the eight-arm radial maze.

The mechanism by which NC-1900, a new pGlu-Asn-Cys(Cys)-Pro-Arg-Gly-NH(2) (AVP(4-9)) analog, improves spatial memory in rats using an eight-arm radial maze was examined. Even at very low doses (0.2 ng/kg for s.c., 1 microg/kg for p.o., 1 fg for i.c.v.) NC-1900 improved scopolamine-induced impairment of spatial memory. NC-1900 (1 ng/kg, s.c.) also improved impairment of spatial memory induced by pirenzepine, a muscarinic(1) (M(1)) receptor antagonist, and by KN-62, a Ca2+/calmodulin (CaM)-dependent protein kinase II inhibitor. [Pmp(1), Tyr(Me)(2)]-Arg(8)-vasopressin, a vasopressin(1A) (V(1A)) receptor antagonist, and nicardipine, L-type Ca2+ blocker, but not OPC-31260, a V(2) antagonist, suppressed the effect of NC-1900 on scopolamine-induced impairment of spatial memory. A microdialysis study showed that NC-1900 did not affect acetylcholine release in the ventral hippocampus (VH) of intact rats or of scopolamine-treated rats. NC-1900 (1 microM) increased [Ca2+](i) in the VH than in the dorsal hippocampus (DH). Pretreatment with nicardipine (1 microM) and Ca2+ -free conditions inhibited the NC-1900-induced [Ca2+](i) response in the VH. Whereas co-administration of NC-1900 (1 microM) and carbachol (500 microM) increased [Ca2+](i) in the VH. Moreover, nicardipine concentration-dependently inhibited the increase in [Ca2+](i) induced by the co-administration of NC-1900 and carbachol in the VH. These results suggest that NC-1900 activates the V(1A) receptor at the postsynaptic cholinergic nerve, and causes a transient influx of intracellular Ca2+ through L-type Ca2+ channels, to interact with the M(1) receptor. The activation of these Ca2+ -dependent processes induced by NC-1900 may be involved in the positive effect of NC-1900 on scopolamine-induced impairment of spatial memory.

Expression of interleukin-6, interleukin-6 receptor (gp80), and the receptor's signal-transducing subunit (gp130) in human normal pituitary glands and pituitary adenomas.

Interleukin-6 (IL-6) is an important cytokine in cell proliferation and differentiation in several organs. It has also been reported that IL-6 plays a role in secretion or release of pituitary hormones in pituitary hormone-secreting cells and pituitary adenomas, but convincing data in situ have not yet been reported. In this study, we examined the participation of IL-6 in the production of pituitary hormones and the differences between human normal pituitary glands and pituitary adenomas by determination of the localization or expression of IL-6, IL-6 receptor (IL-6R, gp80), and the signal-transducing subunit (gp130) of the receptor using immunohistochemical staining and RT-PCR. IL-6 was mainly expressed in ACTH- and FSH/LH-secreting cells in normal pituitary glands, as shown by double staining. gp 80 and gp130 were coexpressed in almost all GH- and PRL-secreting cells and in approximately 30% of FSH/LH-secreting cells. RT-PCR showed that IL-6 mRNA was expressed in only one of all the pituitary adenomas examined, whereas gp 80 and gp 130 mRNAs were detected in all these pituitary adenomas. In conclusion, IL-6 was mainly expressed in ACTH- and FSH/LH-secreting cells, and the receptors were expressed in GH-, PRL- and FSH/LH-secreting cells in human normal pituitary glands. Furthermore, our data emphasized that the mechanism of IL-6 function in human pituitary adenoma cells is distinct from that in normal pituitary cells.

Role of dopaminergic system in core part of nucleus accumbens in hyperlocomotion and rearing induced by MK-801 in rats: a behavioral and in vivo microdialysis study.

We investigated modification of the MK-801 effect on motor activity and extracellular amines concentration by 6-hydroxydopamine (6-OHDA)-induced lesion of core nucleus accumbens (cACC) of rats. In vivo microdialysis-HPLC showed that the concentrations (fmol/microl) of dopamine (DA), dihydroxyphenylacetic acid (DOPAC) and serotonin were 0.738 +/- 0.135, 155.34 +/- 41.01 and 0.334 +/- 0.024, respectively, in the cACC of intact rats. The DOPAC/DA ratio was 264.24 +/- 94.01. Unilateral lesion of the cACC with 6-OHDA (8 microg/microl) substantially reduced DA (-93%) and DOPAC (-97%) in desipramine (30 mg/kg, i.p.)-pretreated rats (6-OHDA+DMI rats) as compared to the 65% reduction rate of both amines in saline-pretreated rats (6-OHDA+saline rats). Moreover, DOPAC was reduced by 72% in 6-OHDA+DMI rats. MK-801 increased DOPAC (426-467%) and DOPAC/DA ratio (180-230%) in intact rats. On the other hand, MK-801 increased DA by 154% and 505% in 6-OHDA+saline and 6-OHDA+DMI rats, respectively. 6-OHDA reduced the effect of MK-801 on DOPAC and DOPAC/DA ratio. In the behavioral studies, MK-801 (0.01-0.3 mg/kg, i.p.) increased locomotor activity and rearing of intact rats. Bilateral 6-OHDA+DMI lesion of the cACC caused greater reduction in the effect of MK-801 (0.1 mg/kg) than that of the shell nucleus accumbens. These results suggest that increased extracellular DOPAC concentration (but not DA) and DOPAC/DA ratio in the cACC plays an important role in MK-801-hyperactivity.

Characteristics of learning and memory impairment induced by delta9-tetrahydrocannabinol in rats.

We investigated the characteristics of delta9-tetrahydrocannabinol (THC)-induced impairment of learning and memory using an 8-arm radial maze task, a water maze, a visual discrimination task with 2 figures and a passive avoidance test in rats. THC (6 mg/kg, i.p.) impaired spatial memory in the standard task of the 8-arm radial maze. THC (4-6 mg/kg, i.p.) selectively impaired working memory in a reference and working memory task of the 8-arm radial maze. Even at a dose of 10 mg/kg, THC did not impair spatial memory in the water maze. In addition, THC at a dose of 6 mg/kg, which had inhibitory effects in the 8-arm radial maze, did not affect performance in the visual discrimination task. These results indicate that at low doses (2-6 mg/kg), THC may not produce visual function abnormalities. THC impaired retrieval (6 mg/kg, i.p.) as well as acquisition (10 mg/kg, i.p.) in the passive avoidance test. The consolidation process was also impaired by i.c.v. injection (100 microg), but not i.p. injection (6-10 mg/kg) of THC. These results suggest that THC-induced impairment of spatial memory is based on the selective impairment of working memory through its effects on acquisition and retrieval processes.

The scopolamine-induced impairment of spatial cognition parallels the acetylcholine release in the ventral hippocampus in rats.

We investigated the relationship between the induction of spatial cognition impairment in the 8-arm radial maze task and regional changes (ventral hippocampus (VH), dorsal hippocampus, frontal cortex, and basolateral amygdala nucleus) in brain acetylcholine (ACh) release using microdialysis in rats treated with muscarinic (M) receptor antagonists. In a behavioral study, two M1 antagonists, scopolamine (0.5 mg/kg, i.p. and 20 microg, i.c.v.) and pirenzepine (80 microg, i.c.v.), but not an M2 antagonist, AF-DX116 (40-80 microg, i.c.v.), disrupted spatial cognition in the 8-arm radial maze task. In brain microdialysis with Ringer's solution containing 0.1 mM eserine sulfate, scopolamine and AF-DX116, but not pirenzepine, increased ACh release in the VH. Moreover, in the bilateral injection of scopolamine (2 microg/side), the VH and dorsomedial thalamus nucleus were important regions for scopolamine-induced impairment of spatial cognition. A simultaneous determination of the behavioral changes revealed that scopolamine (0.5 mg/kg, i.p.) markedly decreased the ACh contents and also increased the ACh release in all regions tested. Especially, the changes in the ACh release of the VH closely paralleled the induction of the scopolamine-induced impairment of spatial cognition. These results suggest that the blocking balance between M1 and M2 muscarinic receptor in the VH therefore plays a major role in the spatial cognition impairment induced by scopolamine in the 8-arm radial maze task.