Occurrence of distinctive cells and effects of irradiance on vascular formation in leaves of shade-tolerant C4 grass Paspalum conjugatum.

The denser leaf vasculature of C4 plants than of C3 plants may suit rapid export of assimilates associated with their higher photosynthetic rate. However, some C4 grasses have partially reduced leaf vasculature with vascular bundle (VB)-free bundle-sheath cells called distinctive cells (DCs). The shade-tolerant C4 grass Paspalum conjugatum has such a reduced leaf vascular system with DCs. We examined whether irradiance during growth affects vascular formation in leaves of P. conjugatum grown under 100%, 30%, or 14% sunlight for 1 month alongside the C4 grass maize. Under all conditions, P. conjugatum leaves had partially reduced vasculature: DCs and incomplete small VBs without phloem occurred between VBs with a normal structure consisting of both xylem and phloem. Shaded plants had less phloem in the small VBs than the full-sunlit plants. In maize, however, all VBs always had both xylem and phloem under all light conditions. The net photosynthetic rate of both grasses was reduced under shade; that of P. conjugatum was always lower than that of maize under all light conditions, but was reduced less by shade than that of maize. The light compensation point was lower in P. conjugatum than in maize, indicating that P. conjugatum acclimatizes better to low light. The reduction of phloem in VBs of P. conjugatum may be an acclimatization to shade, because dense vasculature may be expensive for C4 plants growing in environments where the higher photosynthetic rate is not realized.

Salt stress induces Kranz anatomy and expression of C4 photosynthetic enzymes in the amphibious sedge Eleocharis vivipara.

Eleocharis vivipara Link is a unique amphibious leafless plant of the Cyperaceae. The terrestrial form develops culms with Kranz anatomy and C4-like traits, while the submerged form does culms with non-Kranz anatomy and C3 traits. The submerged form develops new culms with C4-like mode when exposed to air or exogenous abscisic acid. In this study, we investigated whether salt stress (0.05-0.3 M NaCl) has a similar effect. When the submerged form was grown for one month in solutions of 0.1 M NaCl and more, culm growth was strongly suppressed. However, these plants slowly developed new culms that had Kranz anatomy with chloroplast-abundant Kranz bundle sheath cells. Although the culms of the submerged form had only few stomata, culms grown in the NaCl solution had many stomata. The NaCl-grown culms also accumulated large amounts of C4 photosynthetic enzymes (phosphoenolpyruvate carboxylase and pyruvate Pi dikinase), and the cellular localization patterns of these enzymes and ribulose 1,5-bisphosphate carboxylase/oxygenase were similar to those in terrestrial culms. Accumulation of C4 enzymes increased in mature culms of the submerged form (with non-Kranz anatomy) when exposed to 0.2 M NaCl solution for one week. These results suggest that salt stress induces development of Kranz anatomy and expression of C4 photosynthetic enzymes in the submerged C3 form of E. vivipara, whereas the anatomical and biochemical traits of C4 photosynthesis appear to be regulated independently.

Photosynthesis and leaf structure of F1 hybrids between Cymbidium ensifolium (C3) and C. bicolor subsp. pubescens (CAM).

BACKGROUND AND AIMS: The introduction of crassulacean acid metabolism (CAM) into C3 crops has been considered as a means of improving water-use efficiency. In this study, we investigated photosynthetic and leaf structural traits in F1 hybrids between Cymbidium ensifolium (female C3 parent) and C. bicolor subsp. pubescens (male CAM parent) of the Orchidaceae. METHODS: Seven F1 hybrids produced through artificial pollination and in vitro culture were grown in a greenhouse with the parent plants. Structural, biochemical, and physiological traits involved in CAM in their leaves were investigated. KEY RESULTS: Cymbidium ensifolium accumulated very low levels of malate without diel fluctuation, whereas C. bicolor subsp. pubescens showed nocturnal accumulation and diurnal consumption of malate. The F1s also accumulated malate at night, but much less than C. bicolor subsp. pubescens. This feature was consistent with low nocturnal fixation of atmospheric CO2 in the F1s. δ 13C values of the F1s were intermediate between those of the parents. The leaf thickness was thicker in C. bicolor subsp. pubescens than in C. ensifolium, and those of the F1s were more similar to that of C. ensifolium. This was due to the difference in mesophyll cell size. The chloroplast coverage of mesophyll cell perimeter adjacent to intercellular air spaces of C. bicolor subsp. pubescens was lower than that of C. ensifolium, and those of the F1s were intermediate between them. Interestingly, one F1 had structural and physiological traits more similar to those of C. bicolor subsp. pubescens than the other F1s. Nevertheless, all F1s contained intermediate levels of phosphoenolpyruvate carboxylase but as much pyruvate,Pi dikinase as C. bicolor subsp. pubescens. CONCLUSIONS: CAM traits were intricately inherited in the F1 hybrids, the level of CAM expression varied widely among F1 plants, and the CAM traits examined were not necessarily co-ordinately transmitted to the F1s.

Effects of growth temperature and nitrogen nutrition on expression of C3-C4 intermediate traits in Chenopodium album.

Proto-Kranz plants represent an initial phase in the evolution from C3 to C3-C4 intermediate to C4 plants. The ecological and adaptive aspects of C3-C4 plants would provide an important clue to understand the evolution of C3-C4 plants. We investigated whether growth temperature and nitrogen (N) nutrition influence the expression of C3-C4 traits in Chenopodium album (proto-Kranz) in comparison with Chenopodium quinoa (C3). Plants were grown during 5 weeks at 20 or 30 °C under standard or low N supply levels (referred to as 20SN, 20LN, 30SN, and 30LN). Net photosynthetic rate and leaf N content were higher in 20SN and 30SN plants than in 20LN and 30LN plants of C. album but did not differ among growth conditions in C. quinoa. The CO2 compensation point (Γ) of C. album was lowest in 30LN plants (36 µmol mol-1), highest in 20SN plants (51 µmol mol-1), and intermediate in 20LN and 30SN plants, whereas Γ of C. quinoa did not differ among the growth conditions (51-52 µmol mol-1). The anatomical structure of leaves was not considerably affected by growth conditions in either species. However, ultrastructural observations in C. album showed that the number of mitochondria per mesophyll or bundle sheath (BS) cell was lower in 20LN and 30LN plants than in 20SN and 30SN plants. Immunohistochemical observations revealed that lower accumulation level of P-protein of glycine decarboxylase (GDC-P) in mesophyll mitochondria than in BS mitochondria is the major factor causing the decrease in Γ values in C. album plants grown under low N supply and high temperature. These results suggest that high growth temperature and low N supply lead to the expression of C3-C4 traits (the reduction of Γ) in the proto-Kranz plants of C. album through the regulation of GDC-P expression.

Gibberellic acid induces non-Kranz anatomy with C4-like biochemical traits in the amphibious sedge Eleocharis vivipara.

MAIN CONCLUSION: Gibberellic acid induces photosynthetic tissues with non-Kranz anatomy and C4-like biochemical traits in terrestrial-form plants of Eleocharis vivipara. This suggests that the structural and biochemical traits are independently regulated. The amphibious leafless sedge, Eleocharis vivipara Link, develops culms (photosynthetic organs) with C4-like traits and Kranz anatomy under terrestrial conditions, and C3 traits and non-Kranz anatomy under submerged conditions. The conversion from C3 mode to C4-like mode in E. vivipara is reportedly mediated by abscisic acid. Here, we investigated the effects of gibberellic acid (GA) on the differentiation of anatomical and photosynthetic traits because GA is involved in heterophylly in aquatic plants. When 100 µM GA was sprayed on terrestrial plants, the newly developed culms had non-Kranz anatomy in the basal part and Kranz-like anatomy in the upper part. In the basal part, the mesophyll cells were well developed, whereas the Kranz (bundle sheath) cells were reduced and contained few chloroplasts and mitochondria. Stomatal frequency was lower in the basal part than in the upper part. Nevertheless, these tissues had abundant accumulation and high activities of C4 photosynthetic enzymes and had C4-like δ13C values, as seen in the culms of the terrestrial form. When submerged plants were grown under water containing GA-biosynthesis inhibitors (uniconazole or paclobutrazol), the new culms had Kranz anatomy. The culms developed under paclobutrazol had the C3 pattern of cellular accumulation of photosynthetic enzymes. These data suggest that GA induces production of photosynthetic tissues with non-Kranz anatomy in terrestrial plants of E. vivipara, without concomitant expression of C3 biochemical traits. The data also suggest that the differentiation of C4 structural and biochemical traits is regulated independently.

Coleataenia prionitis, a C4-like species in the Poaceae.

C4-like plants represent the penultimate stage of evolution from C3 to C4 plants. Although Coleataenia prionitis (formerly Panicum prionitis) has been described as a C4 plant, its leaf anatomy and gas exchange traits suggest that it may be a C4-like plant. Here, we reexamined the leaf structure and biochemical and physiological traits of photosynthesis in this grass. The large vascular bundles were surrounded by two layers of bundle sheath (BS): a colorless outer BS and a chloroplast-rich inner BS. Small vascular bundles, which generally had a single BS layer with various vascular structures, also occurred throughout the mesophyll together with BS cells not associated with vascular tissue. The mesophyll cells did not show a radial arrangement typical of Kranz anatomy. These features suggest that the leaf anatomy of C. prionitis is on the evolutionary pathway to a complete C4 Kranz type. Phosphoenolpyruvate carboxylase (PEPC) and pyruvate, Pi dikinase occurred in the mesophyll and outer BS. Glycine decarboxylase was confined to the inner BS. Ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) accumulated in the mesophyll and both BSs. C. prionitis had biochemical traits of NADP-malic enzyme type, whereas its gas exchange traits were close to those of C4-like intermediate plants rather than C4 plants. A gas exchange study with a PEPC inhibitor suggested that Rubisco in the mesophyll could fix atmospheric CO2. These data demonstrate that C. prionitis is not a true C4 plant but should be considered as a C4-like plant.

Correction to: Transition from C3 to proto-Kranz to C3-C4 intermediate type in the genus Chenopodium (Chenopodiaceae).

The article Transition from C3 to Correspondence t.

Structural and photosynthetic re-acclimation to low light in C4 maize leaves that developed under high light.

BACKGROUND AND AIMS: C4 plants have higher photosynthetic capacity than C3 plants, but this advantage comes at an energetic cost that is problematic under low light. In the crop canopy, lower leaves first develop under high light but later experience low light because of mutual shading. To explore the re-acclimation of C4 leaves to low light, we investigated the structural and physiological changes of the leaves of maize plants grown in shaded pots. METHODS: Plants were first grown under high light, and then some of them were shaded (20 % of sunlight) for 3 weeks. Four types of leaves were examined: new leaves that developed under low light during shading (L), new leaves that developed under high light (H), mature leaves that developed under high light before shading and were then subjected to low light (H-L) and mature leaves that always experienced high light (H-H). KEY RESULTS: The leaf mass per area, nitrogen and chlorophyll contents per unit leaf area, chlorophyll a/b ratio and activities of C3 and C4 photosynthetic enzymes were lower in H-L than in H-H leaves and in L than in H leaves. Unlike L leaves, H-L leaves maintained the thickness and framework of the Kranz anatomy of H leaves, but chloroplast contents in H-L leaves were reduced. This reduction of chloroplast contents was achieved mainly by reducing the size of chloroplasts. Although grana of mesophyll chloroplasts were more developed in L leaves than in H leaves, there were no differences between H-L and H-H leaves. The light curves of photosynthesis in H-L and L leaves were very similar and showed traits of shade leaves. CONCLUSIONS: Mature maize leaves that developed under high light re-acclimate to low-light environments by adjusting their biochemical traits and chloroplast contents to resemble shade leaves but maintain the anatomical framework of sun leaves.

Transition from C3 to proto-Kranz to C3-C4 intermediate type in the genus Chenopodium (Chenopodiaceae).

The Chenopodiaceae is one of the families including C4 species among eudicots. In this family, the genus Chenopodium is considered to include only C3 species. However, we report here a transition from C3 photosynthesis to proto-Kranz to C3-C4 intermediate type in Chenopodium. We investigated leaf anatomical and photosynthetic traits of 15 species, of which 8 species showed non-Kranz anatomy and a CO2 compensation point (Γ) typical of C3 plants. However, 5 species showed proto-Kranz anatomy and a C3-like Γ, whereas C. strictum showed leaf anatomy and a Γ typical of C3-C4 intermediates. Chenopodium album accessions examined included both proto-Kranz and C3-C4 intermediate types, depending on locality. Glycine decarboxylase, a key photorespiratory enzyme that is involved in the decarboxylation of glycine, was located predominantly in the mesophyll (M) cells of C3 species, in both M and bundle-sheath (BS) cells in proto-Kranz species, and exclusively in BS cells in C3-C4 intermediate species. The M/BS tissue area ratio, number of chloroplasts and mitochondria per BS cell, distribution of these organelles to the centripetal region of BS cells, the degree of inner positioning (vacuolar side of chloroplasts) of mitochondria in M cells, and the size of BS mitochondria also changed with the change in glycine decarboxylase localization. All Chenopodium species examined were C3-like regarding activities and amounts of C3 and C4 photosynthetic enzymes and δ13C values, suggesting that these species perform photosynthesis without contribution of the C4 cycle. This study demonstrates that Chenopodium is not a C3 genus and is valuable for studying evolution of C3-C4 intermediates.

Intracellular position of mitochondria in mesophyll cells differs between C3 and C4 grasses.

In C3 plants, part of the CO2 fixed during photosynthesis in chloroplasts is released from mitochondria during photorespiration by decarboxylation of glycine via glycine decarboxylase (GDC), thereby reducing photosynthetic efficiency. The apparent positioning of most mitochondria in the interior (vacuole side of chloroplasts) of mesophyll cells in C3 grasses would increase the efficiency of refixation of CO2 released from mitochondria by ribulose 1,5-bisphosphate carboxylase/​oxygenase (Rubisco) in chloroplasts. Therefore, in mesophyll cells of C4 grasses, which lack both GDC and Rubisco, the mitochondria ought not to be positioned the same way as in C3 mesophyll cells. To test this hypothesis, we investigated the intracellular position of mitochondria in mesophyll cells of 14 C4 grasses of different C4 subtypes and subfamilies (Chloridoideae, Micrairoideae, and Panicoideae) and a C3-C4 intermediate grass, Steinchisma hians, under an electron microscope. In C4 mesophyll cells, most mitochondria were positioned adjacent to the cell wall, which clearly differs from the positioning in C3 mesophyll cells. In S. hians mesophyll cells, the positioning was similar to that in C3 cells. These results suggest that the mitochondrial positioning in C4 mesophyll cells reflects the absence of both GDC and Rubisco in the mesophyll cells and the high activity of phosphoenolpyruvate carboxylase. In contrast, the relationship between the mitochondrial positioning and enzyme distribution in S. hians is complex, but the positioning may be related to the capture of respiratory CO2 by Rubisco. Our study provides new possible insight into the physiological role of mitochondrial positioning in photosynthetic cells.

Dorsoventral asymmetry of photosynthesis and photoinhibition in flag leaves of two rice cultivars that differ in nitrogen response and leaf angle.

Rice is believed to show photosynthetic symmetry between adaxial and abaxial leaf sides. To verify this, we re-examined dorsoventral asymmetry in photosynthesis, chlorophyll fluorescence and anatomical traits in flag leaves of two Oryza sativa cultivars that differ in nitrogen (N) response and in leaf angle: 'Akenohoshi', a cultivar that can adapt to low-N (LN), with low leaf angle (more erect leaves), and 'Shirobeniya', a cultivar that is unable to adapt to LN, with higher leaf angle. Plants were grown under standard-N (SN) and LN conditions. LN leaves of both cultivars became more erect than SN, but LN Akenohoshi still had more erect ones than Shirobeniya. Contrary to results of previous studies, leaves of both cultivars showed an asymmetry in photosynthetic rate between adaxial and abaxial sides (higher on the adaxial side) under SN. SN leaves of both cultivars showed lower susceptibility to photoinhibition on the adaxial side than on the abaxial side. However, leaves of Akenohoshi showed less asymmetry in these traits under LN than under SN, whereas leaves of Shirobeniya had similar degrees of asymmetry in these traits under both SN and LN. Both cultivars also showed dorsoventral asymmetry in anatomical traits of mesophyll tissue regardless of N level, but the degree of asymmetry was lower in LN Akenohoshi. These data reveal that rice leaves exhibit dorsoventral asymmetry in photosynthetic and anatomical features, and that the degree of asymmetry varies with cultivar and N level. It is suggested that lower leaf angles (particularly in Akenohoshi) in the presence of LN represent a light acclimation to prevent photoinhibition.

Structure and immunocytochemical localization of photosynthetic enzymes in the lamina joint and sheath pulvinus of the C4 grass Arundinella hirta.

The C(4) grass Arundinella hirta exhibits a unique C(4) anatomy, with isolated Kranz cells (distinctive cells) and C(4)-type expression of photosynthetic enzymes in the leaf sheath and stem as well as in the leaf blade. The border zones between these organs are pale green. Those between the leaf blade and sheath and between the sheath and stem are called the lamina joint and sheath pulvinus, respectively, and are involved in gravity sensing. We investigated the structure and localization of C(3) and C(4) photosynthetic enzymes in these tissues. In both zones the epidermis lacked stomata. The inner tissue was composed of parenchyma cells and vascular bundles. The parenchyma cells were densely packed with small intercellular spaces and contained granal chloroplasts with large starch grains. No C(4)-type cellular differentiation was recognized. Western blot analysis showed that the lamina joint and pulvinus accumulated substantial amounts of phosphoenolpyruvate carboxylase (PEPC), pyruvate,Pi dikinase (PPDK), and ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco). Immunogold electron microscopy revealed PEPC in the cytosol and both PPDK and rubisco in the chloroplasts of parenchyma cells, suggesting the occurrence of C(3) and C(4) enzymes within a single type of chlorenchyma cell. These data indicate that the lamina joint and pulvinus have unique expression patterns of C(3) and C(4) enzymes, unlike those in C(4)-type anatomy.

Positive effect of inaudible high-frequency components of sounds on glucose tolerance: a quasi-experimental crossover study.

Although stress significantly impacts on various metabolic syndromes, including diabetes mellitus, most stress management techniques are based on psychological and subjective approaches. This study examined how the presence or absence of the inaudible high-frequency component (HFC) of sounds, which activates deep-brain structures, affects glucose tolerance in healthy participants using the oral glucose tolerance test (OGTT). Sounds containing HFC suppressed the increase in glucose levels measured by incremental area under the curve in the OGTT compared with the otherwise same sounds without HFC. The suppression effect of HFC was more prominent in the older age group and the group with high HbA1c. This suggests that sounds with HFC are more effective in improving glucose tolerance in individuals at a higher risk of glucose intolerance.

Effectiveness and Safety of Ureteroscopic Lithotripsy in Young, Old-Old, and Oldest Old Patients.

Objectives: To investigate the efficacy and safety of ureteroscopic lithotripsy (URSL) in young, old-old, and oldest old patients. Materials and Methods: We enrolled 157 patients who underwent URSL. We divided the patients into young (<65 years), old-old (65-84 years), and oldest old (≥85 years) groups and compared the safety of surgery between young vs old-old and young vs oldest old, separately. Results: In the <65 group, 65 to 85 group, and ≥85 group, respectively, the mean stone diameters were 8.9 ± 4.9, 10.8 ± 7.7, and 11.4 ± 6.3 mm; mean operative time was 76.6 ± 33.1, 86.7 ± 44.7, and 84.0 ± 44.5 min; the stone-free rates were 95.9%, 94.4%, and 96.3%; and the complication rates were 8.2%, 9.8%, and 3.7%. Conclusions: In this study, we showed the feasibility and safety of URSL for elderly people. Although oldest old (≥85 years) people had multiple comorbidities with low performance status, URSL could be performed with acceptable complication rates. Clinical Trial Registration Number: 1809-1.

Structural and biochemical characterization of the C3-C4 intermediate Brassica gravinae and relatives, with particular reference to cellular distribution of Rubisco.

On the basis of its CO(2) compensation concentration, Brassica gravinae Ten. has been reported to be a C(3)-C(4) intermediate. This study investigated the structural and biochemical features of photosynthetic metabolism in B. gravinae. The cellular distribution of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) was also examined in B. gravinae, B. napus L. (C(3)), Raphanus sativus L. (C(3)), and Diplotaxis tenuifolia (L.) DC. (C(3)-C(4)) by immunogold electron microscopy to elucidate Rubisco expression during the evolution from C(3) to C(3)-C(4) intermediate plants. The bundle sheath (BS) cells of B. gravinae contained centrifugally located chloroplasts as well as centripetally located chloroplasts and mitochondria. Glycine decarboxylase P-protein was localized in the BS mitochondria. Brassica gravinae had low C(4) enzyme activities and high activities of Rubisco and photorespiratory enzymes, suggesting that it reduces photorespiratory CO(2) loss by the glycine shuttle. In B. gravinae, the labelling density of Rubisco was higher in the mesophyll chloroplasts than in the BS chloroplasts. A similar cellular pattern was found in other Brassicaceae species. These data demonstrate that, during the evolution from C(3) to C(3)-C(4) intermediate plants, the intercellular pattern of Rubisco expression did not change greatly, although the amount of chloroplasts in the BS cells increased. It also appears that intracellular variation in Rubisco distribution may occur within the BS cells of B. gravinae.

Ammonia emission from rice leaves in relation to photorespiration and genotypic differences in glutamine synthetase activity.

BACKGROUND AND AIMS: Rice (Oryza sativa) plants lose significant amounts of volatile NH(3) from their leaves, but it has not been shown that this is a consequence of photorespiration. Involvement of photorespiration in NH(3) emission and the role of glutamine synthetase (GS) on NH(3) recycling were investigated using two rice cultivars with different GS activities. METHODS: NH(3) emission (AER), and gross photosynthesis (P(G)), transpiration (Tr) and stomatal conductance (g(S)) were measured on leaves of 'Akenohoshi', a cultivar with high GS activity, and 'Kasalath', a cultivar with low GS activity, under different light intensities (200, 500 and 1000 µmol m(-2) s(-1)), leaf temperatures (27·5, 32·5 and 37·5 °C) and atmospheric O(2) concentrations ([O(2)]: 2, 21 and 40 %, corresponding to 20, 210 and 400 mmol mol(-1)). KEY RESULTS: An increase in [O(2)] increased AER in the two cultivars, accompanied by a decrease in P(G) due to enhanced photorespiration, but did not greatly influence Tr and g(S). There were significant positive correlations between AER and photorespiration in both cultivars. Increasing light intensity increased AER, P(G), Tr and g(S) in both cultivars, whereas increasing leaf temperature increased AER and Tr but slightly decreased P(G) and g(S). 'Kasalath' (low GS activity) showed higher AER than 'Akenohoshi' (high GS activity) at high light intensity, leaf temperature and [O(2)]. CONCLUSIONS: Our results demonstrate that photorespiration is strongly involved in NH(3) emission by rice leaves and suggest that differences in AER between cultivars result from their different GS activities, which would result in different capacities for reassimilation of photorespiratory NH(3). The results also suggest that NH(3) emission in rice leaves is not directly controlled by transpiration and stomatal conductance.

Ectopic overexpression of the transcription factor OsGLK1 induces chloroplast development in non-green rice cells.

For systematic and genome-wide analyses of rice gene functions, we took advantage of the full-length cDNA overexpresser (FOX) gene-hunting system and generated >12 000 independent FOX-rice lines from >25 000 rice calli treated with the rice-FOX Agrobacterium library. We found two FOX-rice lines generating green calli on a callus-inducing medium containing 2,4-D, on which wild-type rice calli became ivory yellow. In both lines, OsGLK1 cDNA encoding a GARP transcription factor was ectopically overexpressed. Using rice expression-microarray and northern blot analyses, we found that a large number of nucleus-encoded genes involved in chloroplast functions were highly expressed and transcripts of plastid-encoded genes, psaA, psbA and rbcL, increased in the OsGLK1-FOX calli. Transmission electron microscopy showed the existence of differentiated chloroplasts with grana stacks in OsGLK1-FOX calli cells. However, in darkness, OsGLK1-FOX calli did not show a green color or develop grana stacks. Furthermore, we found developed chloroplasts in vascular bundle and bundle sheath cells of coleoptiles and leaves from OsGLK1-FOX seedlings. The OsGLK1-FOX calli exhibited high photosynthetic activity and were able to grow on sucrose-depleted media, indicating that developed chloroplasts in OsGLK1-FOX rice calli are functional and active. We also observed that the endogenous OsGLK1 mRNA level increased synchronously with the greening of wild-type calli after transfer to plantlet regeneration medium. These results strongly suggest that OsGLK1 regulates chloroplast development under the control of light and phytohormones, and that it is a key regulator of chloroplast development.

Augmentation of Positive Valence System-Focused Cognitive Behavioral Therapy by Inaudible High-Frequency Sounds for Anhedonia: A Trial Protocol for a Pilot Study.

Importance: Recent conceptualizations in Research Domain Criteria have indicated that anhedonia, 1 of 2 core symptoms of depression, which can be treatment resistant, is associated with deficits in the positive valence system, and inaudible high-frequency sound therapy has been shown to enhance reward-related brain circuitry. Hence, cognitive behavioral therapy focusing on the positive valence system enhanced with sound therapy could have a synergistic effect on anhedonia. Objective: To test the augmentation effect of inaudible high-frequency sounds on the efficacy of positive valence system-focused cognitive behavioral therapy to treat anhedonia. Design, Setting, and Participants: In this individual-level allocation, exploratory, single-center randomized superiority pilot trial, patients, therapists, and evaluators will be masked to intervention or placebo assignment. The trial will take place at a national psychiatric referral hospital in Tokyo, Japan, among 44 adult patients with clinically significant anhedonia and moderate to severe depression. Outcomes will be analyzed following the intent-to-treat principle using a repeated-measures mixed model. Intervention: The intervention group will participate in 8 weekly sessions of positive valence system-focused cognitive behavioral therapy with in-session exposure to an inaudible high-frequency sound; the comparison group will undergo cognitive behavioral therapy with in-session exposure to a placebo sound. Main Outcomes and Measures: The primary outcome is anhedonia assessed using the self-reported Snaith-Hamilton Pleasure Scale. The secondary outcome is anhedonia assessed using the clinician-administered version of the Snaith-Hamilton Pleasure Scale. Discussion: Recruitment for this study began in May 2018, and the projected date of final allocation is January 2020. A total of 21 eligible patients were registered for participation as of May 30, 2019. To date, treatments for depression do not guarantee clinically successful outcomes. This pilot trial will provide preliminary evidence of the augmentation effect of high-frequency inaudible sounds on cognitive behavioral therapy for anhedonia. Overall, exposure to an inaudible high-frequency sounds does not require attentional or cognitive effort from either patients or therapists; therefore, results from a future confirmative trial could indicate that cognitive behavioral therapy can be augmented in an effortless manner. Trial Registration: umin.ac.jp/ctr Identifier: UMIN000031948.

Induction of prolonged natural lifespans in mice exposed to acoustic environmental enrichment.

We investigated the effect of acoustic environmental enrichment (EE) on the lifespans and behaviours of mice to the end of their natural lifespan in different acoustic environments. Acoustic EE induced a significantly prolonged natural lifespan (nearly 17% longer) and was associated with increased voluntary movements. However, no correlation between lifespan and voluntary movements was detected, suggesting that increased voluntary movements are not a primary cause of lifespan prolongation. Analyses of individual differences in lifespan demonstrated that lifespan extension induced by acoustic EE could be related to changes in social relationships (e.g., reduction of social conflict) among individuals kept within a cage. Therefore, an acoustic component may be an important factor inducing the positive effects of EE.

A hairy-leaf gene, BLANKET LEAF, of wild Oryza nivara increases photosynthetic water use efficiency in rice.

BACKGROUND: High water use efficiency is essential to water-saving cropping. Morphological traits that affect photosynthetic water use efficiency are not well known. We examined whether leaf hairiness improves photosynthetic water use efficiency in rice. RESULTS: A chromosome segment introgression line (IL-hairy) of wild Oryza nivara (Acc. IRGC105715) with the genetic background of Oryza sativa cultivar 'IR24' had high leaf pubescence (hair). The leaf hairs developed along small vascular bundles. Linkage analysis in BC5F2 and F3 populations showed that the trait was governed by a single gene, designated BLANKET LEAF (BKL), on chromosome 6. IL-hairy plants had a warmer leaf surface in sunlight, probably due to increased boundary layer resistance. They had a lower transpiration rate under moderate and high light intensities, resulting in higher photosynthetic water use efficiency. CONCLUSION: Introgression of BKL on chromosome 6 from O. nivara improved photosynthetic water use efficiency in the genetic background of IR24.