Comprehensive Transcriptomic Analysis of Cordyceps militaris Cultivated on Germinated Soybeans

The ascomycete fungus Cordyceps militaris infects lepidopteran larvae and pupae and forms characteristic fruiting bodies. Owing to its immune-enhancing effects, the fungus has been used as a medicine. For industrial application, this fungus can be grown on geminated soybeans as an alternative protein source. In our study, we performed a comprehensive transcriptomic analysis to identify core gene sets during C. militaris cultivation on germinated soybeans. RNA-Seq technology was applied to the fungal cultures at seven-time points (2, 4, and 7-day and 2, 3, 5, 7-week old cultures) to investigate the global transcriptomic change. We conducted a time-series analysis using a two-step regression strategy and chose 1460 significant genes and assigned them into five clusters. Characterization of each cluster based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases revealed that transcription profiles changed after two weeks of incubation. Gene mapping of cordycepin biosynthesis and isoflavone modification pathways also confirmed that gene expression in the early stage of GSC cultivation is important for these metabolic pathways. Our transcriptomic analysis and selected genes provided a comprehensive molecular basis for the cultivation of C. militaris on germinated soybeans.

Effects of norepinephrine and neuropeptide Y on the contractility of small mesenteric artery from 2K1C and DOCA-salt hypertensive rats

The present study was conducted to investigate the possible role of the sympathetic nervous system in two-kidney, one clip (2K1C) and deoxycorticosterone acetate (DOCA)-salt hypertension. 2K1C and DOCA-salt hypertension were made in Sprague-Dawley rats. Four weeks after induction of hypertension, systolic blood pressure measured in conscious state was significantly higher in 2K1C (216+/-18 mmHg) and DOCA-salt (205+/-29 mmHg) groups than that in control (128+/-4 mmHg). The third branches (<300 micrometer in outer diameter) of the mesenteric artery were isolated and cut into ring segments of 2apprx3 mm in length. Each ring segment was mounted in tissue bath and connected to a force displacement transducer for measurement of isometric tension. The arterial rings were contracted by application of norepinephrine (NE) in a dose-dependent manner. The amplitude of the NE-induced contraction of the vessels was significantly larger in hypertension than in control. The NE-induced contraction was significantly enhanced by neuropeptide Y (NPY) in hypertension. Reciprocally, NPY-elicited vasocontraction was increased by NE in hypertension. These results suggest that the sympathetic nervous system contributes to the development of 2K1C and DOCA-salt hypertension.

Developmental changes of gustatory neurons in nucleus of solitary tract in rats

To learn the developmental changes in intrinsic electrophysiological properties of the second order taste neurons, whole cell recordings from the developing nucleus of the solitary tract neurons were done in brainstem slices of postnatal rats. Rats aged from postnatal 0 to 21 days (P0-P21) were used, being divided into 3 age groups: postnatal first week (P0-P7 days), second week (P8-P14 days), and third week P15-P21 days). Slices containing gustatory NTS were cut horizontally in the thickness of 300 micrometer. Whole cell recordings were obtained from neurons in response to a series of hyperpolarizing and depolarizing current pulses. The intrinsic electrophysiological properties of the rostral NTS (rNTS) neurons were compared among the age groups. Depolarizing current pulses evoked a train of action potentials in all neurons of all age groups. The resting membrane potential and input resistance of the neurons did not show any significant differences during the ostnatal 3 weeks. The time constant, however, decreased during the development. Duration of action potential measured at half maximum amplitude was longer in younger age groups. Both the maximum rate of rise and the maximum rate of fall in the action potential increased during the first 3 weeks postnatal. Electrophysiologically more than half neurons were type III. In summary, it is suggested that developmental changes in electrophysiological properties in rNTS occur during the first three weeks in rats.

Altered Calcium Current of the Vascular Smooth Muscle in Renal Hypertension

The present study was aimed at investigating whether the calcium current in the vascular smooth muscle (VSM) cells is altered in renal hypertension. Two-kidney, one clip (2K1C) and deoxycorticosterone acetate (DOCA)-salt hypertension were made in Sprague-Dawley rats. Rats without clipping the renal artery or implanting DOCA were used as control for 2K1C and DOCA-salt hypertension, respectively. Four weeks after clipping, systolic blood pressure was significantly higher in 2K1C rats than in control (192+/-24 and 119+/-4 mmHg, respectively, n=16 each). DOCA-salt rats also showed a higher blood pressure (180+/-15 mmHg, n=18) compared with control (121+/-6 mmHg, n=14). VSM cells were enzymatically and mechanically isolated from basilar arteries. Single relaxed VSM cells measured 5 ~ 10 mum in width and 70 ~ 150 mum in length were obtained. VSM cells could not be differentiated in size and shape between hypertensive and normotensive rats under light microscopy. High-threshold (L-type) calciumcurrents were recorded using whole-cell patch clamp technique. The amplitude of the current recorded from VSM cells was larger in 2K1C hypertension than in control. Neither the voltage-dependence of the calcium current nor the cell capacitance was significantly affected by 2K1C hypertension. By contrast, the amplitude of the calcium current was not altered in DOCA-salt hypertension. These results suggest that high-threshold calcium current of the VSM cells is altered in 2K1C hypertension, and that calcium channel may not be involved in calcium recruitment of VSM in DOCA-salt hypertension.

Role of endogenous nitric oxide in the control salivary secretion and blood flow

The present study was designed to investigate whether endogenous nitric oxide (EDNO) is involved in submandibular vasodilation and salivation induced by parasympathetic nerve stimulation. Effects of Nw-nitro-L-arginine-methyl ester (L-NAME) which blocks the synthesis of EDNO from L-arginine on the submandibular vasodilation and salivation induced by chorda stimulation or administration of various vasodilators were examined in anesthetized cats. Effect of L-NAME on K+ efflux induced by carbachol was also examined using the excised submandibular slice in vitro. In the submandibular slices, acetylcholine (10(-5) mol/L) or vasoactive intestinal polypeptide (VIP, 10(-5) mol/L) increased NO2 contents, which was prevented by pretreatment with L-NAME. Salivary secretion in response to the chorda stimulation (3 V, 1 msec, 10 ~ 20 Hz) was completely blocked by treatment with atropine (1 mg/kg). Increased blood flow response to the low frequency (1, 2, 5 Hz) stimulation was significantly reduced, whereas the blood flow induced by the higher frequency (10, 20 Hz) stimulation was not affected. Lingual-arterial infusion of L-NAME (100 mg/kg) significantly diminished the vasodilatory and salivary responses to the chorda stimulation at all stimuli frequencies used. Intra-arterial infusion of L-NAME (100 mg/kg) markedly diminished the vasodilatory responses to acetylcholine (5 mug/kg), VIP (5 mug/kg) or bradykinin (5 mug/kg). In the excised submandibular slice, K+ efflux in response to carbachol (10(-5) mol/L) was significantly decrease by pretreatment with L-NAME (10(-5) mol/L). In the isolated submandibular artery precontracted with phenylephrine (10(-5) mol/L), the vasorelaxation induced by ACh (10-7 mol/L) was reversed into a contraction by methylene blue (10(-4) mol/L). These results suggest that EDNO may play an important role in vasodilation and secretion of the submandibular gland.