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Osmoregulation is influenced by a wide variety of biotic and abiotic variables, and maintenance of systemic osmoregulatory homeostasis is critical to insect fitness. Because insects are so small, accurately quantifying renal organ function is technically challenging, and often requires specialized equipment. On top of this, nearly a century of toiling in the laboratory has led to a wide and still growing variety of methods that can be difficult for novice researchers to disentangle. Here, we provide a reference guide for the most used in vitro approaches in the study of insect osmoregulation, including the Ramsay assay, Ussing chamber, epithelial potential measurement, scanning ion-selective electrode technique, and hindgut assays. Along the way, we highlight the history of each methodological innovation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

In the cold, chill susceptible insects lose the ability to regulate ionic and osmotic gradients. This leads to hemolymph hyperkalemia that drives a debilitating loss of cell membrane polarization, triggering cell death pathways and causing organismal injury. Biotic and abiotic factors can modulate insect cold tolerance by impacting the ability to mitigate or prevent this cascade of events. In the present study, we test the combined and isolated effects of dietary manipulations and thermal acclimation on cold tolerance in fruit flies. Specifically, we acclimated adult Drosophila melanogaster to 15 or 25 °C and fed them either a K + -loaded diet or a control diet. We then tested the ability of these flies to recover from and survive a cold exposure, as well as their capacity to protect transmembrane K + gradients, and intracellular Na + concentration. As predicted, cold-exposed flies experienced hemolymph hyperkalemia and cold-acclimated flies had improved cold tolerance due to an improved maintenance of the hemolymph K + concentration at low temperature. Feeding on a high-K + diet improved cold tolerance additively, but paradoxically reduced the ability to maintain extracellular K + concentrations. Cold-acclimation and K + -feeding additively increased the intracellular K + concentration, aiding in maintenance of the transmembrane K + gradient during cold exposure despite cold-induced hemolymph hyperkalemia. There was no effect of acclimation or diet on intracellular Na + concentration. These findings suggest intracellular K + loading and reduced muscle membrane K + sensitivity as mechanisms through which cold-acclimated and K + -fed flies are able to tolerate hemolymph hyperkalemia., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Insect performance is linked to environmental temperature, and surviving through winter represents a key challenge for temperate, alpine and polar species. To overwinter, insects have adapted a range of strategies to become truly cold hardy. However, although the mechanisms underlying the ability to avoid or tolerate freezing have been well studied, little attention has been given to the challenge of maintaining ion homeostasis at frigid temperatures in these species, despite this limiting cold tolerance for insects susceptible to mild chilling. Here, we investigated how prolonged exposure to temperatures just above the supercooling point affects ion balance in freeze-avoidant mountain pine beetle (Dendroctonus ponderosae) larvae in autumn, mid-winter and spring, and related it to organismal recovery times and survival. Hemolymph ion balance was gradually disrupted during the first day of exposure, characterized by hyperkalemia and hyponatremia, after which a plateau was reached and maintained for the rest of the 7-day experiment. The degree of ionoregulatory collapse correlated strongly with recovery times, which followed a similar asymptotical progression. Mortality increased slightly during extensive cold exposures, where hemolymph K+ concentration was highest, and a sigmoidal relationship was found between survival and hyperkalemia. Thus, the cold tolerance of the freeze-avoiding larvae of D. ponderosae appears limited by the ability to prevent ionoregulatory collapse in a manner similar to that of chill-susceptible insects, albeit at much lower temperatures. Based on these results, we propose that a prerequisite for the evolution of insect freeze avoidance may be a convergent or ancestral ability to maintain ion homeostasis during extreme cold stress., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

The physiology of insects is directly influenced by environmental temperature, and thermal tolerance is therefore intrinsically linked to their thermal niche and distribution. Understanding the mechanisms that limit insect thermal tolerance is crucial to predicting biogeography and range shifts. Recent studies on locusts and flies suggest that the critical thermal minimum (CTmin) follows a loss of CNS function via a spreading depolarization. We hypothesized that other insect taxa share this phenomenon. Here, we investigate whether spreading depolarization events occur in butterflies exposed to cold. Supporting our hypothesis, we found that exposure to stressful cold induced spreading depolarization in all 12 species tested. This reinforces the idea that spreading depolarization is a common mechanism underlying the insect CTmin. Furthermore, our results highlight how CNS function is tuned to match the environment of a species. Further research into the physiology underlying spreading depolarization will likely elucidate key mechanisms determining insect thermal tolerance and ecology., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Background and Objectives: Studies analysing the association of albuminuria and prevalent frailty in community-dwelling very old adults are scarce and lack information on incident frailty. We investigated the association of kidney function decline and increase of albuminuria with frailty worsening or death in very old adults., Design: Longitudinal analyses with biennial visits of the Berlin Initiative (cohort) Study and a frailty follow-up of 2.1 years., Setting/subjects: 1,076 participants with a mean age of 84.3 (5.6) years of whom 54% were female., Methods: Partial proportional odds models were used to assess the association of estimated glomerular filtration rate (eGFR) decline and/or albuminuria (albumin creatinine ratio, ACR) with frailty worsening or death., Results: At frailty baseline, 1,076 participants with an eGFR of 50 (13) ml/min/1.73 m2, 48% being prefrail and 31% frail were included. After median 2.1 years, 960 (90%) participants had valid information on frailty transition: 187 (17.5%) worsened and 111 (10.3%) died. In the multivariable model, the odds of frailty worsening for participants with albuminuria in combination with eGFR <60 ml/min/1.73 m2 were elevated [OR (95% CI): 2.47 (1.41-4.31)] compared to participants without albuminuria and eGFR ≥60 ml/min/1.73 m2 as there was a rapid eGFR decline of ≥3 ml/min/1.73 m2 per year [1.55 (1.04-2.33)] and albuminuria trajectories six years prior [1.53 (1.11-2.10)] to frailty baseline. The odds of death for each exposure were even higher., Conclusions: In older adults, advanced stages of CKD and albuminuria alone were associated with 2-fold odds of frailty worsening independent of death., (© The Author(s) 2023. Published by Oxford University Press on behalf of the British Geriatrics Society. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Most insects can acclimate to changes in their thermal environment and counteract temperature effects on neuromuscular function. At the critical thermal minimum, a spreading depolarization (SD) event silences central neurons, but the temperature at which this event occurs can be altered through acclimation. SD is triggered by an inability to maintain ion homeostasis in the extracellular space in the brain and is characterized by a rapid surge in extracellular K+ concentration, implicating ion pump and channel function. Here, we focused on the role of the Na+/K+-ATPase specifically in lowering the SD temperature in cold-acclimated Drosophila melanogaster. After first confirming cold acclimation altered SD onset, we investigated the dependency of the SD event on Na+/K+-ATPase activity by injecting the inhibitor ouabain into the head of the flies to induce SD over a range of temperatures. Latency to SD followed the pattern of a thermal performance curve, but cold acclimation resulted in a left-shift of the curve to an extent similar to its effect on the SD temperature. With Na+/K+-ATPase activity assays and immunoblots, we found that cold-acclimated flies have ion pumps that are less sensitive to temperature, but do not differ in their overall abundance in the brain. Combined, these findings suggest a key role for plasticity in Na+/K+-ATPase thermal sensitivity in maintaining central nervous system function in the cold, and more broadly highlight that a single ion pump can be an important determinant of whether insects can respond to their environment to remain active at low temperatures., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

Hemodiafiltration (HDF), in which both convective and diffusion methods are combined, yields an increased overall solute clearance compared with hemodialysis (HD), specifically for medium and larger molecular weight uremic toxins. Due to uncertainty in the treatment effects, the nephrology community still perceives the implementation of HDF and the achievement of high convective volume as complex. In this article, we review practical aspects of the implementation of HDF that can effectively deliver a high-volume HDF therapy and assure clinical performance to most patients. We also present an overview of the impact of high-volume HDF (compared to HD) on a series of relevant biochemical, patient-reported, and clinical outcomes, including uremic toxin removal, phosphate, Inflammation and oxidative stress, hemodynamic stability, cardiac outcomes, nutritional effects, health-related quality of life, morbidity, and mortality., (© 2022 Wiley Periodicals LLC.)

Background and Objectives: In older adults, data on the age-related course of GFR are scarce, which might lead to misjudgment of the clinical relevance of reduced GFR in old age., Design, Setting, Participants, & Measurements: To describe the course of eGFR in older adults and derive reference values in population-based individuals, we used the longitudinal design of the Berlin Initiative Study (BIS) with a repeated estimation of GFR over a median of 6.1 years of follow-up. In 2069 community-dwelling older individuals (mean inclusion age 80 years, range 70-99), GFR was estimated biennially with the BIS-2 equation, including standardized creatinine and cystatin C levels, sex, and age. We described the crude and adjusted course using a mixed-effects model and analyzed the influence of death on the GFR course applying joint models. GFR slopes were compared using GFR equations on the basis of creatinine and/or cystatin C., Results: We observed a decreasing, thus nonlinear, eGFR decline with increasing age in a population of old adults. The estimated 1-year slope for ages 75 and 90 diminished for men from -1.67 to -0.99 and for women from -1.52 to -0.97. The modeled mean eGFR for men aged ≥79 and women ≥78 was below 60 ml/min per 1.73 m 2 . Multivariable adjustment attenuated slopes only minimally. Taking death into account by applying joint models did not alter the nonlinear eGFR decline. Using eGFR equations on the basis of creatinine only showed linear slope patterns in contrast to nonlinear patterns for equations including cystatin C., Conclusions: The eGFR decline depended on sex and age and changed only marginally after multivariable adjustment but decelerated with increasing age. Equations including cystatin C demonstrated a nonlinear slope challenging the previously assumed linearity of the decline of eGFR in old age., (Copyright © 2022 by the American Society of Nephrology.)

The spotted wing drosophila (SWD), Drosophila suzukii, is a major invasive fruit pest. There is strong consensus that low temperature is among the main drivers of SWD population distribution, and the invasion success of SWD is also linked to its thermal plasticity. Most studies on ectotherm cold tolerance focus on exposure to a single stressful temperature but here we investigated how cold stress intensity affected survival duration across a broad range of low temperatures (-7 to +3 °C). The analysis of Lt 50 at different stressful temperatures (Thermal Death Time curve - TDT) is based on the suggestion that cold injury accumulation rate increases exponentially with the intensity of thermal stress. In accordance with the hypothesis, Lt 50 of SWD decreased exponentially with temperature. Further, comparison of TDT curves from flies acclimated to 15, 19 and 23 °C, respectively, showed an almost full compensation with acclimation such that the temperature required to induce mortality over a fixed time decreased almost 1 °C per °C lowering of acclimation temperature. Importantly, this change in cold tolerance with acclimation was uniform across the range of moderate to intense cold stress exposures examined. To understand if cold stress at moderate and intense exposures affects the same physiological systems we examined how physiological markers/symptoms of chill injury developed at different intensities of the cold stress. Specifically, hsp23 expression and extracellular [K + ] were measured in flies exposed to different intensities of cold stress (-6, -2 and +2 °C) and at various time points corresponding to the same progression of injury (equivalent to 1/3, 2/3 or 3/3 of Lt 50 ). The different cold stress intensities all triggered hsp23 expression following 2 h of recovery, but patterns of expression differed. At the most intense cold stress (-6 and -2 °C) a gradual increase with time was found. In contrast, at +2 °C an initial increase was followed by a dissipating expression. A gradual perturbation of ion balance (hyperkalemia) was also found at all three cold stress intensities examined, with only slight dissimilarities between treatment temperatures. Despite some differences between the three cold intensities examined, the results generally support the hypothesis that intense and moderate cold stress induces the same physiological perturbation. This suggests that cold stress experienced during natural fluctuating conditions is additive and the results also illustrate that the rate of injury accumulation increases dramatically (exponentially) with decreasing temperature (increasing stress)., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)