Balanced Turbo Field Echo with Extended k-space Sampling: A Fast Technique for the Thoracic Ductography.

We evaluated the visibility of the thoracic duct by fast balanced turbo field echo with extended k-space sampling (bTFEe). The thoracic duct of 10 healthy volunteers was scanned by bTFEe using a 1.5-T magnetic resonance imaging (MRI), which was acquired in approximately 2 minutes. Three-dimensional (3D) turbo spin-echo (TSE) was obtained for comparison. The thoracic duct including draining location of the venous system was overall well visualized on bTFEe, compared to TSE.

Optimized 4D time-of-flight MR angiography using saturation pulse.

PURPOSE: To assess arterial visibility on 4D time-of-flight (4D-TOF) by temporal magnetization transfer contrast pulse (t-MTC) and temporal tilted optimized nonsaturating excitation (t-TONE). 3D-TOF magnetic resonance angiography (MRA) is used for the noninvasive assessment of the intracranial arteries. However, it does not provide temporal information for diagnosing hemodynamics. To noninvasively obtain more detailed hemodynamics-related information, we developed a novel time-resolved MRA without the arterial spin labeling technique, termed 4D-TOF MRA using saturation pulse. MATERIALS AND METHODS: On a 3.0T MRI, three techniques were compared to optimize the visibility of the arteries above the circle of Willis; 1) simple 4D-TOF, 2) 4D-TOF with t-MTC, and 3) 4D-TOF with t-MTC and t-TONE. Eight healthy volunteers were scanned with these three sequences. The contrast changes between the background tissue and the arteries in temporal phases were assessed and compared quantitatively and qualitatively. RESULTS: The contrast between the background and the arteries for 4D-TOF with t-MTC and t-TONE was significantly higher than those for the other methods in delayed phases (P < 0.001).The qualitative assessment showed that 4D-TOF with t-MTC and with t-MTC and t-TONE provided better visualization of the intracranial artery than simple 4D-TOF (P < 0.001). CONCLUSION: 4D-TOF with t-TONE and t-MTC enabled observation of the intracranial hemodynamics. Optimized 4D-TOF provides high-quality images without image subtraction, and good visibility of the intracranial arteries even in the prolonged observation time. J. Magn. Reson. Imaging 2016;43:1320-1326.

Dynamic cross-sectional changes of the middle cerebral artery in atherosclerotic stenosis detected by 3·0-Tesla MRI.

OBJECTIVES: Atherosclerotic stenosis of the middle cerebral artery (MCA) is one of the causes of ischemic stroke, but aside from investigations using magnetic resonance angiography (MRA), studies evaluating stenosis are rare. The purpose of this study was to assess dynamic changes of MCA cross section between the systolic and diastolic phases in patients with cerebral infarction using 3·0-Tesla magnetic resonance imaging (3T MRI). METHODS: We assessed 12 stroke patients with M1 stenosis in the MCA and 12 healthy volunteers. We measured MCA cross sections (proximal/distal to stenosis and on the stenosis) in the systolic and diastolic phases by synchronizing imaging with heartbeats, as well as the maximum flow velocity by using cine-phase contrast (PC) MRI. Each patient also underwent conventional MRA. RESULTS: Differences in cross sections between systolic and diastolic phases were significantly smaller in the stenosed artery compared to the distal (P < 0·05) and proximal areas (P < 0·01) in stroke patients. The difference in maximal blood velocity between systolic and diastolic phases at the M1 stenosis was significantly larger than that in the area proximal to the stenosis (P < 0·05). DISCUSSION: We clearly demonstrated dynamic cross-sectional changes in the stenotic areas by 3T MRI, suggesting hemodynamic shear stress, which may further enhance MCA atherosclerosis.

Protein kinase A-dependent substance P expression by pituitary adenylate cyclase-activating polypeptide in rat sensory neuronal cell line ND7/23 cells.

The neurotrophic effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on rat sensory neuronal cell line ND7/23 cells were investigated. PACAP caused a concentration-dependent increase in the number of neurite-bearing cells and the expression of the substance P precursor (PPT) mRNA in 24 h. The effects of PACAP were mimicked by vasoactive intestinal polypeptide with lower potency and dibutyryl-cyclic AMP, and inhibited by inhibitors of protein kinase A, ERK kinase or p38 kinase, KT5720, U0126, or SB203580, respectively. In a PPT promoter luciferase reporter assay, the increase of PPT mRNA was the result of an increase in PPT gene transcriptional activity by PACAP. The increasing effects of PACAP on PPT mRNA were similarly observed in primary cultured rat dorsal root ganglion cells. Thus, PACAP could induce differentiation-like phenomena in sensory neurons in a cAMP-, protein kinase A-, ERK kinase-, and p38 kinase-dependent manner. These results provide evidence of the neurotrophic action of PACAP, which may function to rescue damaged neurons or to switch the neuronal phenotype in injured or inflamed sensory neurons.

[Basic examination of uniformity of image and contrast of multi-transmit system].

In 3.0-T magnetic resonance imaging (MRI), shortened radio frequency (RF) wavelengths cause B(1) inhomogeneity. Multi transmit (MT) has been reported as a method of solving this problem. We compared MT with single transmit (ST) and ST body-tuned CLEAR (BTC) in terms of basic performance because we got an opportunity to use MT. A phantom was used to evaluate the uniformity of the flip angles (FAs) of images, the phantom's diameter, and the specific electric conductivity. To evaluate contrast, volunteers performed the significant difference test, and the changes in the FA of the phantom were measured. MT and BTC were better than ST in terms of the uniformity of the images. MT had the best contrast. The results showed that the uniformity of the images and the contrast were improved using MT compared with ST because MT can be uniformly irradiated to an object using two individual RF transmitters.

A simple and highly sensitive HPLC method with fluorescent detection for determination of pipecolic acid in mouse brain areas.

A simple and highly sensitive high-performance liquid chromatography method for the determination of pipecolic acid in mouse brain areas was developed. After homogenization of brain and pretreatment with o-phthalaldehyde, the pipecolic acid and (2S,3S)-3-methylpyrrolidine-2-carboxylic acid (internal standard) were derivatized with 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride at 70 °C for 15 min in basic medium (pH 9.0). The fluorescent derivatives of pipecolic acid and internal standard were separated on a reversed-phase column by stepwise elution using acetic acid (30 mM)-acetonitrile at 50 °C and detected by fluorescence measurement at 316 nm (excitation) and 403 nm (emission). The detection limit (signal-to-noise ratio=3) of pipecolic acid was 13 fmol per injection. The recovery was about 106.7%. The precision (relative standard deviation) was 3.2%.

The antioxidative and antilipidemic effects of different molecular weight chitosans in metabolic syndrome model rats.

The effect of high and low molecular weight chitosans (HMC; 1000 kDa, LMC; 30 kDa) on oxidative stress and hypercholesterolemia was investigated using male 6-week-old Wistar Kyoto rats as a normal model (Normal-rats) and spontaneously hypertensive rat/ND mcr-cp (SHP/ND) as a metabolic syndrome model (MS-rats), respectively. In Normal-rats, the ingestion of both chitosans over a 4 week period resulted in a significant decrease in total body weight (BW), glucose (Gl), triglyceride (TG), low density lipoprotein (LDL) and serum creatinine (Cre) levels. The ingestion of both chitosans also resulted in a lowered ratio of oxidized to reduced albumin and an increase in total plasma antioxidant activity. In addition to similar results in Normal-rats, the ingestion of only HMC over a 4 week period resulted in a significant decrease in total cholesterol levels in MS-rats. Further, the ingestion of LMC resulted in a significantly higher antioxidant activity than was observed for HMC in both rat models. In in vitro studies, LMC caused a significantly higher reduction in the levels of two stable radicals, compared to HMC, and the effect was both dose- and time-dependent. The findings also show that LDL showed strong binding in the case of HMC. These results suggest that LMC has a high antioxidant activity as well as antilipidemic effects, while HMC results in a significant reduction in the levels of pro-oxidants such as LDL in the gastrointestinal tract, thereby inhibiting the subsequent development of oxidative stress in the systemic circulation in metabolic model rats.

Functional expression of 5-HT2A receptor in osteoblastic MC3T3-E1 cells.

In the previous study, we reported the gene expression for proteins related to the function of 5-hydroxytryptamine (5-HT, serotonin) and elucidated the expression patterns of 5-HT(2) receptor subtypes in mouse osteoblasts. In the present study, we evaluated the possible involvement of 5-HT receptor subtypes and its inactivation system in MC3T3-E1 cells, an osteoblast cell line. DOI, a 5-HT(2A) and 5-HT(2C) receptor selective agonist, as well as 5-HT concentration-dependently increased proliferative activities of MC3T3-E1 cells in their premature period. This effect of 5-HT on cell proliferation were inhibited by ketanserin, a 5-HT(2A) receptor specific antagonist. Moreover, both DOI-induced cell proliferation and phosphorylation of ERK1 and 2 proteins were inhibited by PD98059 and U0126, selective inhibitors of MEK in a concentration-dependent manner. Furthermore, treatment with fluoxetine, a 5-HT specific re-uptake inhibitor which inactivate the function of extracellular 5-HT, significantly increased the proliferative activities of MC3T3-E1 cells in a concentration-dependent manner. Our data indicate that 5-HT fill the role for proliferation of osteoblast cells in their premature period. Notably, 5-HT(2A) receptor may be functionally expressed to regulate mechanisms underlying osteoblast cell proliferation, at least in part, through activation of ERK/MAPK pathways in MC3T3-E1 cells.

Expression of mRNA for 5-HT2 receptors and proteins related to inactivation of 5-HT in mouse osteoblasts.

We analyzed the expression of 5-HT(2) receptors and proteins related to inactivation of 5-HT in primary cultures of mouse osteoblasts. The mRNA for the 5-HT(2A) receptor was detectable in anaplastic osteoblasts as well as in differentiated and matured osteoblasts. The mRNA for the 5-HT(2B) receptor and 5-HT transporter was undetectable in anaplastic osteoblasts and became detectable in differentiated and matured osteoblasts. It was suggested that 5-HT might regulate the proliferation of anaplastic osteoblasts through the 5-HT(2A) receptor without control by 5-HT-inactivating mechanisms. The differentiation and maturation of osteoblasts might be regulated by the activation of the 5-HT(2B) receptor under the control of 5-HT inactivation.

Inhibitory effect of juniperonic acid (Delta-5c,11c,14c,17c-20:4, omega-3) on bombesin-induced proliferation of Swiss 3T3 cells.

Juniperonic acid (Delta-5c,11c,14c,17c-20:4, JA) is a polymethylene-interrupted (PMI) fatty acid that occurs in Biota orientalis. In this study, we found that JA has an antiproliferative activity. Swiss 3T3 cells were preloaded with fatty acids before stimulation with bombesin, a mitogenic neuropeptide, and proliferation of the cells was assessed by [(3)H]thymidine incorporation. Preloading of linoleic acid (Delta-9c,12c-18:2) significantly enhanced bombesin-induced proliferation. In contrast, preloading of eicosapentaenoic acid (Delta-5c,8c,11c,14c,17c-20:5, EPA) suppressed proliferation. Likewise, cells preloaded with JA showed a significantly curtailed response to bombesin. The antiproliferative potency of JA was equivalent to that of EPA. Sciadonic acid (Delta-5c,11c,14c-20:3), an omega-6 analogue of JA did not show antiproliferative activity, suggesting the importance of the omega-3 double bond rather than the PMI structure. The EPA-like activity of JA may be involved in the pharmaceutical activity of biota seeds, a psychoactive Chinese traditional medicine.

Contribution of the peripheral 5-HT 2A receptor to mechanical hyperalgesia in a rat model of neuropathic pain.

We investigated the effect of 5-HT receptor antagonists on mechanical hyperalgesia observed in a neuropathic pain rat model prepared by chronic constriction injury of the sciatic nerve. NAN-190, a 5-HT 1A receptor antagonist, (-)-pindolol, a 5-HT 1A/1B receptor antagonist, and tropisetron, a 5-HT(3/4) receptor antagonist, did not affect the pain threshold in the hyperalgesic hind limb to the same extent as in the normal hind limb. However, sarpogrelate and ketanserin, 5-HT 2A receptor antagonists, significantly elevated the pain threshold in the hyperalgesic hind limb, but not in the normal hind limb. In spite of its high affinity for the 5-HT 2A receptor, methysergide only slightly elevated the pain threshold in the hyperalgesic hind limb. Pre-treatment with methysergide significantly antagonized the inhibitory effect of sarpogrelate on hyperalgesia. Furthermore, the 5-HT 2A receptor specific binding activity of 3H-ketanserin determined for the hyperalgesic hind limb did not differ from that of the normal hind limb. From these results, we propose that the 5-HT 2A receptor in the hyperalgesic hind paw function as an agonist-independent active receptor following constriction of the sciatic nerve, and that sarpogrelate and ketanserin act as inverse agonists of this receptor and suppress its activation. Methysergide may act as a neutral antagonist that blocks the effect of inverse agonists on the 5-HT 2A receptor.

Identification of the binding sites and selectivity of sarpogrelate, a novel 5-HT2 antagonist, to human 5-HT2A, 5-HT2B and 5-HT2C receptor subtypes by molecular modeling.

The aim of the present study was to investigate the binding sites interactions and the selectivity of sarpogrelate to human 5-HT(2) receptor family (5-HT(2A), 5-HT(2B) and 5-HT(2C) receptor subtypes) using molecular modeling. Rhodopsin (RH) crystal structures were used as template to build structural models of the human serotonin-2A and -2C receptors (5-HT(2A)R, 5-HT(2C)R), whereas for 5-HT(2B)R, we used our previously published three-dimensional (3D) models based on bacteriorhodopsin (BR). Sarpogrelate, a novel 5-HT(2)R antagonist, was docked to the receptors. Molecular dynamics (MD) simulations produced the strongest interaction for 5-HT(2A)R/sarpogrelate complex. Upon binding, sarpogrelate constraints aromatic residues network (Trp(3.28), Phe(5.47), Trp(6.48), Phe(6.51), Phe(6.52) in 5-HT(2A)R; Phe(3.35), Phe(6.51), Trp(7.40) in 5-HT(2B)R; Trp(3.28), Phe(3.35), Phe(5.47), Trp(6.48), Phe(6.51), Phe(6.52) in 5-HT(2C)R) in a stacked configuration, preventing activation of the receptor. The models suggest that the structural origin of the selectivity of sarpogrelate to 5-HT(2A)R vs both 5-HT(2B)R and 5-HT(2C)R comes from the following results: (1) The tight interaction between the antagonist and the transmembrane domain (TMD) 3. Asp(3.32) neutralizes the cationic head and interacts simultaneously with carboxylic group hydrogen of the antagonist molecule. (2) Due to steric hindrance, Ser(5.46) (vs Ala(5.46) in 5HT(2B) and 5HT(2C)) prevents sarpogrelate to enter deeply inside the hydrophobic core of the helix bundle and to interact with Pro(5.50). (3) The side chain of Ile(4.56) (vs Ile(4.56) in 5HT(2B)R and Val(4.56) in 5HT(2C)R) constraints sarpogrelate to adjust its position by translating toward the strongly attractive Asp(3.32). These results are in good agreement with binding affinities (pKi) of sarpogrelate for 5-HT(2) receptor family expressed in transfected cell.

5-HT-induced, 5-HT3 receptor-mediated, and ruthenium red- and capsaicin-sensitive positive chronotropic effects in the isolated guinea pig atrium.

We investigated the mechanisms of 5-HT-induced tachycardia, which we reported previously to be triggered by 5-HT3 receptor stimulation, in the isolated guinea pig atrium in comparison with that induced by isoproterenol and histamine. We found that 5-HT-induced tachycardia was completely inhibited by ruthenium red. 5-HT-induced tachycardia was reduced in the capsaicin pre-treated atrium as well as in the presence of capsaicin. The effects of isoproterenol and histamine were not affected by ruthenium red or capsaicin treatment. Furthermore, 5-HT-induced tachycardia was found to be potentiated by thiorphan, an inhibitor of peptide degeneration. Calcitonin gene-related peptide (CGRP) (1-37), a full agonist of CGRP1-like receptors, was found to act selectively as a potent stimulator of chronotropic action. CGRP (8-37), an antagonist of CGRP1-type receptors, inhibited 5-HT-induced tachycardia as well as effects induced by CGRP (1-37). The observation that tetrodotoxin failed to affect 5-HT-induced tachycardia excluded the involvement of 5-hydroxytryptaminergic interneurons. Thus, we confirmed that the mechanism of 5-HT-induced tachycardia is distinct from that induced by isoproterenol and histamine. In conclusion, the activation of 5-HT3 receptors on the sensory nerve terminals brought about ruthenium red-sensitive Ca2+ influx and resulted in the release of CGRP from capsaicin-sensitive stores, and then CGRP stimulated CGRP1-like receptors to produce 5-HT-induced tachycardia.