Search

Given its limited accessibility, the CA2 area has been less investigated compared to other subregions of the hippocampus. While the development of transgenic mice expressing Cre recombinase in the CA2 has revealed unique features of this area, the use of mouse lines has several limitations, such as lack of specificity. Therefore, a specific gene delivery system is required. Here, we confirmed that the AAV-PHP.eB capsid preferably infected CA2 pyramidal cells following retro-orbital injection and demonstrated that the specificity was substantially higher after injection into the lateral ventricle. In addition, a tropism for the CA2 area was observed in organotypic slice cultures. Combined injection into the lateral ventricle and stereotaxic injection into the CA2 area specifically introduced the transgene into CA2 pyramidal cells, enabling us to perform targeted patch-clamp recordings and optogenetic manipulation. These results suggest that AAV-PHP.eB is a versatile tool for specific gene transduction in CA2 pyramidal cells.

Tyramide signal amplification (TSA) is a highly sensitive method for histochemical analysis. Previously, we reported a TSA system, biotinyl tyramine-glucose oxidase (BT-GO), for bright-filed imaging. Here, we develop fluorochromized tyramide-glucose oxidase (FT-GO) as a multiplex fluorescent TSA system. FT-GO involves peroxidase-catalyzed deposition of fluorochromized tyramide (FT) with hydrogen peroxide produced by enzymatic reaction between glucose and glucose oxidase. We showed that FT-GO enhanced immunofluorescence signals while maintaining low background signals. Compared with indirect immunofluorescence detections, FT-GO demonstrated a more widespread distribution of monoaminergic projection systems in mouse and marmoset brains. For multiplex labeling with FT-GO, we quenched antibody-conjugated peroxidase using sodium azide. We applied FT-GO to multiplex fluorescent in situ hybridization, and succeeded in labeling neocortical interneuron subtypes by coupling with immunofluorescence. FT-GO immunofluorescence further increased the detectability of an adeno-associated virus tracer. Given its simplicity and a staining with a high signal-to-noise ratio, FT-GO would provide a versatile platform for histochemical analysis.

Phosphorus (P) is generally considered the most common limiting nutrient for productivity of mature tropical lowland forests growing on highly weathered soils. It is often assumed that P limitation also applies to young tropical forests, but nitrogen (N) losses during land-use change may alter the stoichiometric balance of nutrient cycling processes. In the Amazon basin, about 16% of the original forest area has been cleared, and about 30-50% of cleared land is estimated now to be in some stage of secondary forest succession following agricultural abandonment. Here we use forest age chronosequences to demonstrate that young successional forests growing after agricultural abandonment on highly weathered lowland tropical soils exhibit conservative N-cycling properties much like those of N-limited forests on younger soils in temperate latitudes. As secondary succession progresses, N-cycling properties recover and the dominance of a conservative P cycle typical of mature lowland tropical forests re-emerges. These successional shifts in N:P cycling ratios with forest age provide a mechanistic explanation for initially lower and then gradually increasing soil emissions of the greenhouse gas nitrous oxide (N(2)O). The patterns of N and P cycling during secondary forest succession, demonstrated here over decadal timescales, are similar to N- and P-cycling patterns during primary succession as soils age over thousands and millions of years, thus revealing that N availability in terrestrial ecosystems is ephemeral and can be disrupted by either natural or anthropogenic disturbances at several timescales.

Here we present the within-site, seasonal, and interannual variations of the carbon (δ13C) and nitrogen (δ15N) isotope ratios of leaves, wood, bark and litter from four sites in the Amazon region, Brazil. Samples were collected in Manaus (3° 06′07″ S; 60°01′30″ W), Ji-Parana (10°53′07″ S; 61°57′06″ W), and Santarem (2°26′35″ S; 54°42′30″ W) with mean annual precipitation of 2207, 2040 and 1909 mm respectively. The overall average for all leaf samples was−32.3 ± 2.5‰ for δ13C and +5.8 ± 1.6‰ for δ15N (n = 756). The leaf δ values at these sites were often but not always statistically distinct from each other. The d13C values varied from−37.8‰ to−25.9‰ Pronounced differences in δ13C values occurred with height associated with differences in forest structure. The δ13C of leaf dry matter showed seasonal variations associated with the length of the dry season, despite the fact that total annual precipitation was similar among the studied sites. Leaf δ15N values ranged from+0.9‰ to a maximum value of+10.9‰, and the Santarem sites showed more enriched values than Manaus and Ji-Parana sites. No seasonal variation was detected in the δ15N of leaves, but significant differences were observed among sites and with changes in canopy height. The isotope ratio data are consistent with our current understanding of the roles of light, water availability, and recycling of soil-respired CO2 influences on δ13C and consistent with our understanding that an open nitrogen cycle can lead to high δ15N values despite a significant number of legumes in the vegetation.