Disappearance of perihepatic lymph node enlargement after hepatitis C viral eradication with direct-acting antivirals.

Perihepatic lymph node enlargement (PLNE) which has been shown to be negatively associated with hepatocellular carcinoma (HCC) occurrence is frequently observed in chronic liver disease; however, changes in the state of perihepatic lymph nodes after eradication of hepatitis C virus (HCV) have not been investigated yet. We aimed to evaluate this issue. We enrolled 472 patients with chronic HCV infection who achieved viral eradication with direct-acting antivirals (DAA). We investigated whether the status of perihepatic lymph nodes changed before and after HCV eradication (primary endpoint). We also evaluated the association between PLNE and clinical findings such as liver fibrosis or hepatocellular injury before HCV eradication (secondary endpoint). Perihepatic lymph node enlargement was detected in 164 of 472 (34.7%) patients before DAA treatment. Surprisingly, disappearance of PLNE was observed in 23.8% (39 patients) of all PLNE-positive patients after eradication of HCV. Disappearance of PLNE was not associated with baseline clinical parameters or changing rates of clinical findings before and after DAA treatment. At baseline, presence of PLNE was significantly associated with a lower serum HCV-RNA level (P = .03), a higher serum AST level (P = .004) and a higher ALT level (P < .001) after adjustment for sex and age. In conclusion, PLNEs became undetectable after DAA treatment in 23.8% of PLNE-positive patients. Further study with a longer follow-up period is needed to clarify the clinical importance of this phenomenon especially in relationship with the risk of HCC development.

Trunk muscularity in throwers.

This study aimed to examine and compare the trunk muscularity of track and field throwers and non-athletes, and its predictive value to the physical performance of the athletes. Using a magnetic resonance imaging method, the skeletal muscle volume (SMV) of the trunk (SMV(trunk)) was determined in 19 strength trained athletes and 18 non-athletes. Also, the SMV of upper, middle and lower regions of the trunk was calculated in every 33% of the trunk length. For the athletes, the maximum weight (1RM) of squat, high clean, and deadlift, and shot forward throwing score were measured. The SMV(trunk) in the athletes was 10% greater than that of non-athletes, with a larger difference in the upper region of the trunk. Step-wise multiple regression analysis indicated that the SMV of the lower region was a significant contributor for predicting the 1RM values of the 3 tasks, as well as the shot forward throwing score. The current results indicate that, while the muscularity of the trunk in track and field throwers is characterized by predominant development in the upper region, the muscularity in the lower region is a determinant factor for the 1RM values of the squat, high clean, and deadlift and shot forward throwing score.

Component analysis of dental porcelain for assisting dental identification.

The fluorescence of porcelain crowns recovered from the mouth of an unknown murder victim, and several control porcelain samples, were examined by fluorescent examination lamps. The fluorescence from two of the control samples was quite similar to that from the porcelain crowns recovered from the victim. To increase the objectivity of the results by quantitative analysis, the composition of each porcelain crown and control sample was also evaluated by wave dispersion X-ray microanalyser. The elements detected from the porcelain crowns of the victim matched those of two of the porcelain samples. Later, the antemortem dental records and radiographs of the victim were obtained through a dentist, who had recognized the name of the porcelain manufacturer in a postmortem dental information request placed on the Japanese Dental Association web page. Although component analysis of dental porcelain may be an effective means of assisting dental identification, a more rapid and non-destructive analysis for detecting the elements is required. The energy dispersive X-ray fluorescence (EDXRF) spectrometer was used for a pilot study of identification of porcelain composition.

The liver and the hematolymphoid system: I. The regulation of nylon-passed spleen cell proliferation by active factors released from syngeneic nonparenchymal liver cells.

Nylon-passed spleen cells were found to proliferate when cultured with syngeneic nonparenchymal adherent liver cells and their culture supernatants. The supernatants contained IL-1, IL-6, GM-CSF, and IFN (alpha + beta) activities but not IL-2 and IL-3 activities. The IFN level was higher in early culture sup (2-24 hr) than in later culture sup (48-72 hr). Proliferation was greatly increased by anti-IFN (alpha + beta) serum in the spleen cells cultured in the earlier sup. This antiserum increased the spleen cell proliferation only slightly in the later culture sup. This suggests that nonparenchymal liver cells produce two factors, one having a suppressor, and the other an enhancer action, with IFN being one of the suppressor factors. With culture time, DNA synthesis of spleen cells increased and IL-2 and IL-3 activities were generated in the culture sup. Cells proliferated during culture were found to be morphologically lymphocytes, granulocytes, and macrophages. The mechanisms by which nonparenchymal liver cells regulate the hematolymphoid system are discussed based on our observations.

Vasospasm in monkeys resolves because of loss of and encasement of subarachnoid blood clot.

BACKGROUND AND PURPOSE: We studied in monkeys why vasospasm resolves after subarachnoid hemorrhage (SAH). METHODS: Monkeys underwent angiography and right (n=17) or bilateral (n=8) SAH. Animals with bilateral SAH underwent angiography 1, 3, 5, and 7 days later. Animals with right SAH underwent angiography 7 days later. The clot was then not removed (n=5), removed and replaced with fresh clot (n=7), or removed and not replaced (n=5). At the same time on day 7, the removed clot (n=12) or fresh clot (n=5) was placed on the left side. Angiography was repeated every 2 days until day 14. RESULTS: SAH caused significant vasospasm on day 7 that resolved by day 14. Removal of clot on day 7 resulted in more rapid resolution of vasospasm. Placement of fresh clot onto arteries that had already been exposed to clot for 7 days produced vasospasm that persisted without resolving for an additional 7 days. Placement of 7-day-old clot from the right onto previously unexposed left arteries or of clot from blood removed from an animal 7 days after SAH caused significantly more rapid onset of vasospasm compared with de novo vasospasm. Microscopic examination of the clots showed they were surrounded by macrophages 7 days after SAH. Arterial compliance and contractility were reduced in relation to duration of the exposure of arteries to clot. CONCLUSIONS: Vasospasm resolves because of loss of subarachnoid blood clot. We hypothesize that reduced spasmogen release from the clot contributes to resolution of vasospasm. There was no response in the cerebral arteries that rendered them less responsive to the subarachnoid clot.

An ultrastructural study of nerve profiles in the myenteric plexus of the rabbit colon.

The ultrastructure of the myenteric plexus from the rabbit colon was examined in both conventionally fixed tissue and also material fixed with the chromaffin method. Montages of the ganglia were analysed semi-quantitatively. Six main types of axon profile are described and classified on a morphological consideration of the vesicle population. Most axon types formed synapses with myenteric neurons. Two kinds of chromaffin-positive nerve fibre were seen, one containing a predominance of small granular vesicles, the other containing many flattened vesicles. The difficulties in relating axon profile types to putative transmitters are discussed.

An ultrastructural study of neurons and non-neuronal cells in the myenteric plexus of the rabbit colon.

The myenteric plexus of the rabbit colon showed a degree of structural organization that was unusually high for the peripheral nervous system, providing a basis for the complex integrative activity which is known to occur. It resembled central nervous tissue in several respects: a wide range of neuron types was present; the proportion of glial cells to neurons was about 2:1; and there was a densely packed, avascular neuropil, not penetrated by connective tissue. Most neurons had at least one surface exposed to the extraganglionic space. Clear evidence was obtained for spontaneous neuronal degeneration. Three types of non-neuronal (glial) cells were observed: Type 1, which was most common, contained many 10 nm 'gliofilaments' and resembled enteric glial cells or astrocytes in the central nervous system; Type 2, composing about 5% of the glial cells, had few filaments; Type 3 was seen only rarely, had a small dark nucleus, little cytoplasm, may have been of extraganglionic origin and resembled microglia of the central nervous system. Fibroblast-like cells were also present in extraganglionic sites. Schwann cells could not be identified within the myenteric ganglia.

Long-lasting activation of cation current by low concentration of endothelin-1 in mouse fibroblasts and smooth muscle cells of rabbit aorta.

1. Recombinant human ETA receptors were expressed in a mouse fibroblast cell line (Ltk- cell) and functional coupling of the receptors with Ca2+ permeable channels at low concentrations of endothelin-1 (ET-1) was investigated using whole-cell recordings and monitoring the changes in intracellular free Ca2+ concentrations ([Ca2+]i) with a Ca2+ indicator, fluo-3. A similar type of coupling was investigated in freshly dispersed vascular smooth muscle cells (VSMCs) of rabbit thoracic aorta by use of whole-cell recordings. 2. In Ltk- cells expressing recombinant human ETA receptors, concentrations of ET-1 (10(-8) M, 10(-9) M) evoked an initial transient peak and a subsequent sustained elevation in [Ca2+]i whereas a lower concentration of ET-1 (10(-10) M) evoked only a sustained elevation of [Ca2+]i. After removal of extracellular Ca2+, ET-1 evoked only an initial peak without a sustained elevation of [Ca2+]i. The sustained elevation induced by 10(-10) M ET-1 was blocked by 300 microM mefenamic acid (a cation channel blocker) but not by 10 microM nifedipine (a blocker of voltage-operated Ca2+ channel). 3. In whole-cell recordings with Ltk- cells, a brief (3-5 min) application of ET-1 (10(-10) M) induced a sustained inward current at a holding potential of -60 mV. The current-voltage relationship revealed that the reversal potential of the ET-1-induced current was close to 0 mV (1.9 mV) and was not altered by reducing the concentration of Cl- in the bath solution, indicating that the current is carried by cations. The current was reversibly blocked by 300 microM mefenamic acid, and it persisted after all cations in the bath solution had been replaced by Ca2+ (5 or 30 mM) and nonpermeant cation N-methyl-D glucamine,indicating that the ET-1-activated channel is permeable to Ca2+. Activation of the current was independent of membrane potential and the current was induced even after addition of a high concentration (10 mM) of a Ca2+ chelator, EGTA, to the pipette solution.4. In whole-cell recordings from rabbit aortic VSMCs, ET-l (101-10 M) induced a sustained inward current at a holding potential of -60 mV. The reversal potential was - 12 mV and was not altered when the concentration of Cl- in the pipette solution was decreased, indicating that the current is carried by cations. Again activation of the current was independent of membrane potential and was observed even after addition of a high concentration (10 mM) of a Ca2+ chelator, EGTA to the pipette solution. The current was reversibly blocked by 300 microM mefenamic acid and was permeable to Ca2+,showing marked similarities to ET-1-induced cationic current in Ltk- cells.5. These results indicate that in Ltk- cells transfected with cDNA for recombinant ETA receptors andVSMCs, ETA receptors can functionally couple with a nonselective cation channel permeable to Ca2+.Thus the present data suggest that the cation channel plays an essential role in the sustained elevation of[Ca2+]i at low concentrations of ET-l by causing Ca2+ entry through the channel.

Inhibitory effect of nitrovasodilators and cyclic GMP on ET-1-activated Ca(2+)-permeable nonselective cation channel in rat aortic smooth muscle cells.

1. In single vascular smooth muscle cells (VSMCs) isolated from the aortae of male Wistar rats, we examined the effects of nitric oxide (NO) donors such as sodium nitroprusside (SNP) and S-nitroso-N-acetyl-DL-penicillamine (SNAP), and 8-bromo-guanosine-3':5'-cyclic monophosphate (8-bromo-cyclic GMP) on endothelin-1 (ET-1)-activated Ca(2+)-permeable nonselective cation channel by use of whole-cell recordings of patch-clamp technique and monitoring of intracellular free Ca(2+)-concentration ([Ca2+]i) with fura-2 real-time digital microfluorometry. 2. ET-1 evoked an initial transient peak and a subsequent sustained elevation in [Ca2+]i. After removal of extracellular Ca2+. ET-1 evoked only an initial transient peak without a sustained phase. Nifedipine (1 microM), a specific blocker of the L-type voltage-operated Ca2+ channel (VOC), reduced the sustained phase to about 40% of the control level. The remaining part of the sustained phase was abolished by 30 microM SK&F 96365, a blocker of nonselective cation channels. 3. The nifedipine-resistant sustained elevation in [Ca2+]i was abolished by 100 microM SNP, 10 microM SNAP and 300 microM 8-bromo-cyclic GMP. Neither SNP, SNAP nor 8-bromo-cyclic GMP significantly affected the basal level of [Ca2+]i. 4. In a VSMC clamped at a holding potential of -60 mV with K+ in the pipette solution replaced by Cs+, application of 10(-8) M ET-1 induced an inward current with an increase in baseline fluctuation. With fluctuation analysis, unit conductance of the ET-1-induced current was calculated to be about 21 pS. The ET-1-induced current was linearly related to the membrane potentials with its reversal potential of -5.5 mV. 5. The ET-1-induced current was reversibly and completely inhibited by 30 microM SK&F 96365 or 500 microM Cd2+. The current inhibited by SK&F 96365 or Cd2+ was linearly related to membrane potential with a reversal potential of about -5 mV. 6. The ET-1-induced current was reversibly and completely inhibited by 100 microM SNP, 10 microM SNAP and 300 microM 8-bromo-cyclic GMP. The current inhibited by SNP, SNAP or 8-bromo-cyclic GMP showed linear voltage-dependence and reversed at about -5 mV. 7. In a bath solution in which all cations were replaced by 30 mM Ca2+ and 100 mM nonpermeant cation N-methyl-D-glucamine (NMDG), ET-1 evoked a current with a reversal potential of -11 mV, from which PCa2+/Pcs1 was calculated to be 2.1. This Ca2+ current was also abolished by 100 microM SNP, 10 microM SNAP and 300 microM 8-bromo-cyclic GMP. The current inhibited by SNP, SNAP or 8-bromo-cyclic GMP showed linear voltage-dependence and reversed at about -11 mV. 8. These results taken together indicate that NO through a cyclic GMP signalling pathway inhibits ET-1-activated Ca(2+)-permeable nonselective cation channels, thereby suppressing the sustained increase in [Ca2+]i. Thus, the present study indicates that this Ca(2+)-permeable nonselective cation channel is an important target for nitrovasodilators.

Pharmacological characterization of Ca2+ entry channels in endothelin-1-induced contraction of rat aorta using LOE 908 and SK&F 96365.

We have recently shown that endothelin-1 (ET-1) activates two types of Ca2+-permeable nonselective cation channels (designated NSCC-1 and NSCC-2) and store-operated Ca2+ channel (SOCC). These channels can be pharmacologically discriminated using Ca2+ channel blockers such as SK&F 96365 and LOE 908. Here we characterized Ca2+ entry channels involved in ET-1-induced contractions of rat thoracic aortic rings and increases in the intracellular free Ca2+ concentration ([Ca2+]i) of single smooth muscle cells using these blockers. LOE 908 or a blocker of voltage-operated Ca2+ channel nifedipine had no effect on the contractions and increases in [Ca2+]i induced by thapsigargin or ionomycin, whereas SK&F 96365 abolished them. The contractions and increases in [Ca2+]i induced by ET-1 depended on extracellular Ca2+ but were resistant to nifedipine. The responses to lower concentrations (< or =0.1 nM) of ET-1 were abolished by either SK&F 96365 or LOE 908. The responses to higher concentrations (> or = 1 nM) were abolished by SK&F 96365, but were partially resistant to LOE 908. SK&F 96365 inhibited the LOE 908-resistant contractions induced by higher concentrations of ET-1 with IC50 values similar to those for contractions induced by thapsigargin or ionomycin. These results show that the contractions and increases in [Ca2+]i of rat aortic smooth muscles at lower concentrations of ET-1 involve only one Ca2+ entry channel which is sensitive to SK&F 96365 and LOE 908 (NSCC-2), whereas those at higher concentrations of ET-1 involve another Ca2+ entry channel which is sensitive to SK&F 96365 but resistant to LOE 908 (SOCC) in addition to the former channel.

The involvement of a novel mechanism distinct from the thrombin receptor in the vasocontraction induced by trypsin.

1. The vasocontracting effect of a serine protease trypsin and its mechanisms were investigated by monitoring the isometric tension in endothelium-denuded rings of rabbit thoracic aortae and its effects on intracellular free Ca2+ concentrations ([Ca2+]i) in dispersed rabbit vascular smooth muscle cells with a Ca2+ indicator fura-2. The actions of trypsin were compared with those of thrombin. 2. Both thrombin and trypsin reversibly contracted aortic rings without endothelium in a concentration-dependent manner. The vasocontraction induced by trypsin was well correlated with the protease activity of trypsin actually added to the tissue baths containing the aortic rings and was completely blocked by soybean trypsin inhibitor and phenylmethylsulphonyl fluoride (PMSF), a serine protease inhibitor. 3. The trypsin-induced contractions of the aortic rings were not the result of irreversible damage to vascular smooth muscle cells, since the contractile responses induced by noradrenaline or 30 mM KCl were unaffected by pretreatment with trypsin. 4. The contractions induced by either thrombin or trypsin were reduced to about 30% of control responses after removal of extracellular Ca2+, indicating that most of the contraction is dependent on extracellular Ca2+. By contrast, the contractions induced by either of the proteases were reduced by an antagonist of L-type voltage-operated Ca2+ channels, nifedipine, to about 70% of control responses, indicating that both nifedipine-sensitive and -resistant Ca2+ channels are involved in these contractions. 5. In the aortic rings precontracted by a maximally effective concentration of thrombin, the second application of thrombin virtually failed to induce contractions but trypsin could still induce contractions amounting to 10% of control values by it's protease activity. 6. After the first application of a maximal concentration of thrombin, the second application of thrombin could not induce an increase in [Ca2+]i, but an application of trypsin could still induce an increase in [Ca2+]i in dispersed rabbit vascular smooth muscle cells. 7. These data suggest that in addition to activation of a thrombin receptor, trypsin can contract rabbit aortae by a proteinase-activated receptor 2 or a novel mechanism.

Identification of the interstitial cells of Cajal.

Observation of whole mount stretch preparations using the zinc-iodide-osmic acid method reveals a wide variety of interstitial cells in different tissue layers of the guinea-pig small intestine. And a subsequent electron-microscopic examination and survey of references makes clear that the interstitial cells of Cajal (ICC) depicted in original drawings of Cajal are heterogeneous and correspond to different types of interstitial cells. The myenteric ICC are characterized by long dichotomous branching processes which constitute cellular networks independent from the nerve plexus and form many gap junctions at their tips. Their ultrastructure is similar to that of fibroblasts and they have no basal lamina. The myenteric ICC show strong immunoreactivity for vimentin and the c-kit receptor, and probably correspond to the intestinal pacemaker cells. Within the circular muscle layer, ICC are represented by the cells that are closely associated with fine nerve bundles. The ICC have various shapes, ranging from bipolar to stellate, depending on the running pattern of the nerve fibers that they are associated with. They show fibroblast-like ultrastructure and have no basal lamina. They form gap junctions with smooth muscle cells and are immunoreactive for vimentin. On the other hand, ICC associated with the deep muscular plexus described in the guinea-pig by Cajal could not be clearly identified. However, it is suggested that the ICC in this location may correspond to glycogen-rich cells possessing a basal lamina. Although they show a fairly well-developed rough endoplasmic reticulum, Golgi apparatus and immunoreactivity for vimentin, ICC of the deep muscular plexus are probably specialized smooth muscle cells in nature.

Comparative morphology of interstitial cells of Cajal: ultrastructural characterization.

The shape, distribution, and ultrastructural features of interstitial cells of Cajal (ICC) of different tissue layers and organs of the rat and guinea-pig digestive tract were described and compared with the corresponding cells in other species including mice, dogs, and humans, as reported in the literature. By light microscopy, the best marker for ICC appeared to be immunoreactivity for c-Kit. Ultrastructurally, ICC were characterized by the presence of many mitochondria, bundles of intermediate filaments, and gap junctions, which linked ICC with each other. However, ICC were morphologically heterogeneous and had particular features, depending on their tissue and organ location and species. ICC in the deep muscular plexus of the small intestine and in the submuscular plexus of the colon were the most like smooth muscle cells, and had a distinct basal lamina and numerous caveolae. In contrast, ICC of Auerbach's plexus at all levels of the gastrointestinal tract were the least like smooth muscle cells. They most closely resembled unremarkable fibroblasts. ICC within the circular muscle layer were intermediate in form. In addition to the tissue specificity, some organ and species specificity could be distinguished. The structural differences between ICC may be determined by their microenvironment, including the effects of mechanical force, type of nerve supply, and spacial relationship with smooth muscle cells.

Comparison of R- and S-form lipopolysaccharides fractionated from Escherichia coli UKT-B lipopolysaccharide in pyrogen and Limulus tests.

Different LPS was shown to have a relatively different proportion of O-specific chain-less (R-form) LPS by polyacrylamide gel electrophoresis (PAGE) with sodium deoxycholate (DOC). By using DOC-PAGE, S-form LPS having O-chain with approximately 11 repeating units on average (S-Fr) and O-chain-less LPS (R-Fr) were separated from Escherichia coli UKT-B S-form LPS. Significantly stronger pyrogenicity was observed in R-Fr than in S-Fr when measured on the weight basis. Similar result was observed in Limulus test. Comparing biological activities of different S-form LPS, attention should be given to the amounts of co-existing R-form LPS.

Role of nonselective cation channels as Ca2+ entry pathway in endothelin-1-induced contraction and their suppression by nitric oxide.

The present study was carried out to clarify the role of nonselective cation channels as a Ca2+ entry pathway in the contraction and the increase in [Ca2+]i induced by endothelin- in endothelium-denuded rat thoracic aorta rings, and their suppression by nitric oxide (NO). In Ca2+-free medium, the endothelin-1-induced contraction was suppressed to about 20% of control values, although the increase in [Ca2+]i became negligible. The contraction and the increase in [Ca2+]i monitored by fura 2 fluorescence were unaffected by a blocker of L-type voltage-operated Ca2+ channels nifedipine. A blocker of nonselective cation channels 1-[beta-[3-(4-methoxyphenyl)propoxyl]-4-methoxyphenethyl]-1H-imida zole . HCl(SK&F 96365) suppressed the endothelin-1-induced contraction and increase in [Ca2+]i to the level similar to that after removal of extracellular Ca2+. SK&F 96365 had no further effect on the endothelin-1-induced contraction in the absence of extracellular Ca2+. The endothelin-1-induced contraction and increase in [Ca2+]i were abolished by a donor of NO sodium nitroprusside. The effects of another NO donor 3-morpholinosydnonimine (SIN-1) were also tested and yielded essentially similar results to those for sodium nitroprusside on the endothelin-1-induced contraction. Furthermore, the inhibitory effects of sodium nitroprusside could be blocked with a guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ) at 30 microM. These findings suggest that Ca2+ entry through nonselective cation channels but not voltage-operated Ca2+ channels plays a critical role in the endothelin-1-induced increase in [Ca2+]i and the resulting contraction and that inhibition by NO of the endothelin-1-induced contraction is mainly the result of blockade of Ca2+ entry through these channels.

Three-dimensional visualization of the intraganglionic structures of the rat myenteric plexus: scanning electron microscopy with the connective tissue digestion method.

The rat myenteric plexus was chemically microdissected and the internal structures of the ganglia were demonstrated under a scanning electron microscope. The present preparation offers a view of the three-dimensional features of ganglion cells and permits their surface structures and the size and pattern of varicosities to be observed. Numerous finger-like processes were observed on the cell bodies of the studded neurons in close relationship with the varicose axons. The intramuscular branches of the plexus were also microdissected and their running pattern was observed.

Direct contacts between Auerbach's ganglia and smooth muscle cells in the small intestine of the rat.

The present electron microscopic study demonstrated direct contacts between Auerbach's ganglia and longitudinal smooth muscle cells in the rat small intestine. The muscle cells were often observed to extend small, foot-like processes to the Auerbach's ganglia. These processes were in contact with glial cells in the ganglia without an intervening basal lamina, or were in contact with intraganglionic axon varicosities containing many synaptic vesicles. The processes in contact with glial cells may anchor the muscle cells to the ganglia during muscle contraction and those in contact with axon varicosities may function as synaptic sites between ganglion and longitudinal muscle cells.

The vasorelaxation of cerebral arteries by carbon monoxide.

Carbon monoxide (CO) is known to increase cerebral blood flow, but the effect of CO on the vascular tone of large cerebral arteries is uncertain. We tested whether CO affects cerebral artery tone by measuring tension generated by ex vivo segments of dog basilar artery upon exposure to CO. In cerebral artery segments contracted with either KCl or prostaglandin F(2alpha), CO caused a concentration-related relaxation beginning with a concentration of 57 microM. Relaxation did not occur if CO was administered in the presence of bubbling carboxygen (95% O(2):5% CO(2)), which reduces greater than 99% of CO from the solution. Furthermore, the CO-induced relaxation of cerebral artery segments was reduced in the presence of the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 microM)or the potassium channel blocker tetraethylammonium (TEA, 1 mM). Neither ODQ nor TEA completely eliminated the relaxation caused by CO and there was no additive effect if ODQ and TEA were administered together. These results suggest that cerebral arteries are directly relaxed by CO and that this relaxation depends upon the activation of guanylyl cyclase and the opening of potassium channels.

High-speed liquid chromatographic determination of antiepileptic drugs in human plasma.

A high-speed liquid chromatographic method for the simultaneous determinations of diphenylhydantoin, phenobarbital and carbamazepine in human blood plasma is presented. This method involves two step extraction procedures with chloroform and uses 2 X 50 cm long stainless steel columns packed with a anion exchange resin, with a mobile phase of 4 mM ammonium phosphate buffer solution, pH 6.2 at a flow rate of 0.40 ml/min. The results presented show linear calibration curves and quantitative determinations as low as 1.0 mug of each drug added to 0.5 ml plasma. This method has a sensitivity sufficient to detect human plasma levels after therapeutic clinical doses.

Re-evaluation of fibroblasts and fibroblast-like cells.

Scanning electron microscopic observations of connective tissue cells show a new aspect of the nature of fibroblasts, and the subsequent broad survey of references makes clear that fibroblasts of many tissues have various features which are regarded as atypical of fibroblasts, and at the same time that various connective tissue cells in different organs have features typical of fibroblasts. Both morphological and functional features of fibroblasts are more or less common to those of fibroblast-like cells, and differences among these cells are quantitative rather than qualitative. Therefore, it is almost impossible to set clear-cut criteria for distinguishing genuine fibroblasts from a large population of fibroblast-like cells. The majority of cells sharing features of fibroblasts, if not all, seem to belong to the same population of cells. They are probably adapted to special functional needs in their own micro-environment that are peculiar to local or pathological or experimental conditions. It is proposed to categorize these cells into subtypes depending on their main functions: 1, fibrogenesis; 2, tissue skeleton or barrier; 3, intercellular communication system; 4, gentle contractile machinery; 5, endocrine activity; and 6, vitamin A-storing. Re-evaluation of fibroblasts and fibroblast-like cells is required to facilitate their better understanding.