Proton-driven sodium secretion in a saline water animal.

Aquatic animals residing in saline habitats either allow extracellular sodium concentration to conform to environmental values or regulate sodium to lower levels. The latter strategy requires an energy-driven process to move sodium against a large concentration gradient to eliminate excess sodium that diffuses into the animal. Previous studies of invertebrate and vertebrate species indicate a sodium pump, Na+/K+ ATPase, powers sodium secretion. We provide the first functional evidence of a saline-water animal, Aedes taeniorhynchus mosquito larva, utilizing a proton pump to power this process. Vacuolar-type H+ ATPase (VHA) protein is highly expressed on the apical membrane of the posterior rectal cells, and in situ sodium flux across this epithelium increases significantly in larvae held in higher salinity and is sensitive to Bafilomycin A1, an inhibitor of VHA. We also report the first evidence of splice variants of the sodium/proton exchanger, NHE3, with both high and low molecular weight variants highly expressed on the apical membrane of the posterior rectal cells. Evidence of NHE3 function was indicated with in situ sodium transport significantly inhibited by a NHE3 antagonist, S3226. We propose that the outward proton pumping by VHA establishes a favourable electromotive gradient to drive sodium secretion via NHE3 thus producing a hyperosmotic, sodium-rich urine. This H+- driven Na+ secretion process is the primary mechanism of ion regulation in salt-tolerant culicine mosquito species and was first investigated over 80 years ago.

Biocultural and social determinants of ill health and early mortality in a New Mexican paediatric autopsy sample.

RESULTS.: Hispanic children have higher odds of growth stunting than non-Hispanic White children. Native American children die younger and have higher odds of respiratory diseases and porous lesions than Hispanic and non-Hispanic Whites. Rural/urban location does not significantly impact age at death, but housing type does. Individuals who lived in trailers/mobile homes had earlier ages at death. When intersections between housing type and housing location are considered, children who were poor and from impoverished areas lived longer than those who were poor from relatively well-off areas. CONCLUSIONS.: Children's health is shaped by factors outside their control. The children included in this study embodied experiences of social and ELS and did not survive to adulthood. They provide the most sobering example of the harm that social factors (structural racism/discrimination, socioeconomic, and political structures) can inflict.

Gene knockout of NHX-3 in the soil nematode Caenorhabditis elegans leads to broad-spectrum compensatory regulation of Na+/H+ exchangers, antiporters, and the V-type H+-ATPase.

Na+/H+ exchangers are directly involved in a variety of an animal's essential physiological processes such as ionoregulation, acid-base regulation, nitrogenous waste excretion, and nutrient absorption. While nine NHX isoforms have been identified in Caenorhabditis elegans, the physiological importance of each isoform is not understood. The current study aimed to further our knowledge of NHX-3 which has previously been suggested to be involved in the movement of ammonia and acid-base equivalents across the nematode's hypodermis. Although NHX-3 knockout mutant nematodes exported H+ and imported Na+ at slower rates than wild-type nematodes, attempts to inhibit the NHX activity of mutant nematodes using amiloride and EIPA caused an unexpected increase in hypodermal H+ export and did not impact Na+ fluxes suggesting that the different H+ and Na+ transport profiles of the nematodes are likely due to compensatory changes in the mutants in response to the NHX-3 knockout, rather than the loss of NHX-3's physiological function. Significant changes in the mRNA expression of 7 other NHX isoforms, 2 Na+/H+ antiporter isoforms, and the V-type H+-ATPase were detected between wild-type and mutant nematodes. Furthermore, mutant nematodes possessed significantly reduced rates of cytochrome C oxidase activity and ammonia excretion rates, indicating the knockout of NHX-3 induced fundamental changes in metabolism that could impact the nematode's need to eliminate metabolic end-products like H+ and ammonia that relate to NHX transport. While C. elegans is a popular genetic model with cheap and accessible commercial mutants, our findings suggest caution in interpretation of results in studies using mutants to study physiological traits and the biological significance of specific transporters.

Transport of tetraethylammonium by the Malpighian tubules of Trichoplusia ni: Regional specialization and the influence of diet.

Insect Malpighian tubules (MTs) play a major role in elimination of many potentially toxic compounds, including the organic cation tetraethylammonium (TEA). This paper examines transport of TEA by different segments of the MTs of the cabbage looper, Trichoplusia ni. The results show that the proximal ileac plexus (PIP) region of the MTs plays a dominant role in secretion of the organic cation TEA and that the rate of secretion is altered by feeding; principal cells of the proximal ileac plexus in tubules from larvae with full guts secreted TEA at higher rates than did the same cells in tubules of larvae in which the gut was empty. Michaelis-Menten analysis revealed that TEA secretion by the PIP was saturable and was blocked in a concentration-dependent manner by the organic cation cimetidine. For larvae reared from eggs on TEA-rich diet, higher concentrations of TEA in fluid secreted by the ileac plexus of tubules, and lower concentrations of TEA in the hemolymph, relative to larvae reared on control diet, is consistent with an upregulation of TEA transport in response to higher levels of dietary intake of an exogenous organic cation. The distal and proximal regions of the ileac plexus were also differentiated on the basis of transepithelial and basolateral membrane potentials and the influence of these electrical potentials on organic cation transport are discussed.

Skeletal manifestations of disease experience: Length of illness and porous cranial lesion formation in a contemporary juvenile mortality sample.

OBJECTIVES: Porous lesions of the orbit (cribra orbitalia [CO]) and cranial vault (porotic hyperostosis [PH]) are used as skeletal indicators of childhood stress. Because they are understudied in contemporary populations, their relationship to disease experience is poorly understood. This paper examines the relationship between length of childhood illness and CO/PH formation in a clinically documented sample. "Turning points," which identify the window for lesion formation for CO/PH, are defined, implications for hidden heterogeneity in frailty are considered. METHODS: Data are from 333 (199 males; 134 females) pediatric postmortem computed tomography scans. Individuals died in New Mexico (2011-2019) and are 0.5 to 15.99 years (mean = 7.1). Length of illness was estimated using information from autopsy and field reports. Logistic regression was used to estimate predicted probabilities, odds ratios, and the temporal window for lesion formation. RESULTS: Illness, single bouts, or cumulative episodes lasting over 1 month is associated with higher odds of CO; individuals who were never sick have lower odds of having PH. This relationship was consistent for fatal and incidental illnesses that did not cause death. The developmental window for CO formation appears to close at 8 years. CONCLUSIONS: Those ill for over 1 month are more likely to have CO/PH than those with acute illnesses. Some individuals lived sufficiently long to form CO/PH but died of illness. Others with lesions died of circumstances unrelated to disease. This indicates hidden variation in robusticity even among ill individuals with CO/PH, which is vital in interpreting lesion frequencies in the archeological record.

A perspective on insect water balance.

Insects have a large ratio of surface area to volume because of their small size; thus, they face the potential for desiccation in the terrestrial environment. Nonetheless, they constitute over half of identified species and their success on land can be attributed, in part, to adaptations that limit water loss and allow for effective gains of water from food, fluids or atmospheric water vapour. Reduction of water loss from the gut involves sophisticated mechanisms of ion recycling and water recovery by epithelia of the Malpighian tubules and hindgut. Water loss across the body surface is greatly reduced by the evolution of very thin but highly impermeable lipid-rich layers in the epicuticle. Respiratory water loss can be reduced through effective spiracular control mechanisms and by mechanisms for convective rather than diffusive gas exchange. In addition to extracting water from food sources, some insects are capable of absorption of atmospheric water vapour through processes that have evolved independently in multiple groups.

Blending physiology and RNAseq to provide new insights into regulation of epithelial transport: switching between ion secretion and reabsorption.

This Review addresses the means by which epithelia change the direction of vectorial ion transport. Recent studies have revealed that insect Malpighian (renal) tubules can switch from secreting to reabsorbing K+. When the gut of larval lepidopterans is empty (during the moult cycle) or when the larvae are reared on K+-deficient diet, the distal ileac plexus segment of the tubule secretes K+ from the haemolymph into the tubule lumen. By contrast, in larvae reared on K+-rich diet, ions and fluid are reabsorbed from the rectal lumen into the perinephric space surrounding the cryptonephridial tubules of the rectal complex. Ions and fluid are then transported from the perinephric space into the lumen of the cryptonephridial tubules, thus supplying the free segments of the tubule downstream. Under these conditions, some of the K+ and water in the tubule lumen is reabsorbed across the cells of the distal ileac plexus, allowing for expansion of haemolymph volume in the rapidly growing larvae, as well as recycling of K+ and base equivalents. RNA sequencing data reveal large-scale changes in gene transcription that are associated with the switch between ion secretion and ion reabsorption by the distal ileac plexus. An unexpected finding is the presence of voltage-gated, ligand-gated and mechanosensitive ion channels, normally seen in excitable cells, in Malpighian tubules. Transcriptomic surveys indicate that these types of channels are also present in multiple other types of vertebrate and invertebrate epithelia, suggesting that they may play novel roles in epithelial cell signalling and regulation of epithelial ion transport.

Vacuolar H+-ATPase and Na+/K+-ATPase energize Na+ uptake mechanisms in the nuchal organ of the hyperregulating freshwater crustacean Daphnia magna.

The nuchal organ of the embryos and neonates of the cladoceran, Daphnia magna, has been shown to be a site of Na+ influx and H+, NH4+ and Cl- efflux. This study combines the scanning ion-selective electrode technique with application of inhibitors of specific transporters to assess the mechanisms of Na+ transport across the nuchal organ. Na+ influx across the nuchal organ was inhibited both by inhibitors of the Na+/K+-ATPase (ouabain, bufalin) and by inhibitors of the vacuolar H+-ATPase (bafilomycin, N-ethylmaleimde, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole, KM91104, S-nitrosoglutathione). Na+ influx was unaffected by the epithelial Na+ channel blocker benzamil, but was sensitive to ethylisopropyl amiloride and elevated external ammonium concentrations, consistent with roles for Na+/H+ and Na+/NH4+ exchangers in the apical membrane but not Na+ channels. Transport across the basolateral membrane into the haemolymph is proposed to involve the Na+/K+-ATPase and a thiazide-sensitive Na+/Cl- cotransporter.

Novel mechanisms of epithelial ion transport: insights from the cryptonephridial system of lepidopteran larvae.

Lepidopterans are among the most widespread and easily recognized insects. Whereas adult lepidopterans are known for their beauty and ecological importance as pollinators and sources of food for other animals, larvae are economically important pests of forests and agricultural crops. In the larval body, rapid growth while feeding on plant-based diet is associated with extreme alkalinity (up to pH = 11) of the midgut lumen that helps digest plant proteins. Additionally, the presence of plant secondary metabolites which serve as anti-herbivory agents requires uninterrupted excretory function, accomplished primarily by the Malpighian tubules (MTs). The so-called cryptonephridial condition, along with extreme regional heterogeneity of the MTs, and the ability to rapidly and reversibly alter the direction of epithelial ion transport are features that allow uninterrupted MT functioning and recycling of base equivalents. Studies of MTs in lepidopteran larvae have revealed that rapid adjustments in epithelial ion transport include unexpected roles for voltage-gated, ligand-gated and mechanosensitive ion channels, as well as gap junctions. These molecular components are present in epithelia of a variety of vertebrates and invertebrates and thus are likely to constitute a universal epithelial toolkit for rapid autonomous regulation of epithelial function.

Voltage-gated calcium channels regulate K+ transport in the Malpighian tubules of the larval cabbage looper, Trichoplusia ni.

Transporting epithelia are tissues that specialize in the directional movements of ions and water and are typically either secretory or reabsorptive. Recent work on the Malpighian tubule of larval lepidopterans (caterpillars) demonstrated that the distal ileac plexus segment of this epithelium is capable of rapidly switching between ion secretion and reabsorption. Subsequent transcriptomic studies suggested expression of voltage-gated ion channels in the lepidopteran MTs (which are not contractile and not innervated). The present study shows that isolated MTs of larval Trichoplusia ni express α1, ß2, and α2δ4 subunits of voltage-gated Ca2+ channel CaV1 and that pan-CaVα immunoreactivity is present in the apical and basolateral membranes of the principal cells. Basolateral membrane potential (Vbl) in isolated MTs of larval Trichoplusia ni was influenced by CaV1 functioning; pharmacological inhibition of CaV1 reversed Vbl from inside-negative to inside-positive, and also reduced transepithelial potential (Vte), lowered [Ca2+]i and reversed the direction of K+ transport from secretion to reabsorption. Thus, our findings indicate that a functional CaV1 channel is necessary for constitutive K+ secretion observed in isolated preparations of lepidopteran MTs. Lastly, Vte and Vbl of isolated MTs were influenced by changes in bathing saline [K+]. Our findings suggest that epithelia may rely on CaV channels to enable robust ion secretion and downregulation of CaV channels, together with other transcriptional changes, enables ion reabsorption.

Mechanisms of nitrogen excretion in insects.

Avoiding the toxic effects of ammonia derived from catabolism of proteins and nucleic acids typically involves synthesis of the less soluble compound uric acid in insects, although some species which are not water stressed excrete ammonia directly. Some dipterans metabolize uric acid further to allantoin or urea. Uric acid plays diverse roles as a nitrogenous waste, nitrogen store, pigment, antioxidant and possibly a signaling molecule. Multiple transporters are implicated in urate transport, including members of the ABC and SLC families. Excretion of ammonia by the Malpighian tubules, hindgut, or anal papillae involves multiple transporters, including Na+/K+-ATPase, Rhesus glycoproteins, ammonia transporters (AMTs) and possibly a hyperpolarization-activated cyclic nucleotide-gated K+ channel (HCN).

Mechanisms and regulation of chloride transport in the Malpighian tubules of the larval cabbage looper Trichoplusia ni.

Malpighian tubules (MTs) and the hindgut together constitute the excretory system of insects. Larvae of lepidopterans (butterflies and moths) demonstrate the so-called cryptonephric arrangement, where the distal blind end of each MT is embedded into the rectal complex. The rest of the free tubule is modified into several distinct regions that differ greatly in the transport of cations and water. However, relatively little is known about the transport of counter-anions (e.g., Cl- and HCO3-) by the MTs of lepidopteran larvae. In the current study we used ion-selective microelectrodes to characterize Cl- transport in the distinct regions of the free MT of the larval Trichoplusia ni. Firstly, we note that Cl- transport in the MTs is sensitive to the Cl- concentration of the bathing saline, and several regions of the MTs are capable of either secreting or reabsorbing Cl-. In the distal ileac plexus (DIP), a region previously characterized by cellular heterogeneity and its ability to switch between cation secretion and reabsorption, principal cells (PCs) toggled between Cl- reabsorption (in high-Cl- saline) and Cl- secretion (in low-Cl- saline). In contrast, secondary cells (SCs) in the DIP secreted Cl- regardless of saline Cl- concentration. Mechanistically, we have detected a number of 'leak' and ligand-gated Cl- channels (ClC) and demonstrated that Cl- channels are involved in Cl- secretion. Additionally, we demonstrated that the lumen-positive transepithelial potential increased in response to glycine. Using the scanning ion-selective electrode technique we demonstrated that glycine stimulated Cl- secretion by SCs, but not by PCs. In contrast, when MTs were deprived of glycine, a decrease in Cl- secretion, coupled with a decrease in the TEP, was observed. In contrast to the effects of glycine, an active dose of helicokinin reduced Cl- secretion by PCs, but not by SCs. Lastly, we detected expression of chloride-bicarbonate exchangers (CBE) in all regions of the free tubule. Scans of H+ transport across the tubule indicated that base equivalents are likely reabsorbed across the ileac plexus. Blocking ClC or CBE led to secretion of a more basic fluid, indicating lack of base reabsorption. We suggest that the transport of Cl- in the MTs of larval lepidopterans (i) may be correlated with the reabsorption of base, (ii) may be sensitive to Cl- concentration in the haemolymph, and (iii) could be regulated by helicokinin and glycine.

Only as strong as the weakest link: structural analysis of the combined effects of elevated temperature and pCO2 on mussel attachment.

Predicting how combinations of stressors will affect failure risk is a key challenge for the field of ecomechanics and, more generally, ecophysiology. Environmental conditions often influence the manufacture and durability of biomaterials, inducing structural failure that potentially compromises organismal reproduction, growth, and survival. Species known for tight linkages between structural integrity and survival include bivalve mussels, which produce numerous byssal threads to attach to hard substrate. Among the current environmental threats to marine organisms are ocean warming and acidification. Elevated pCO2 exposure is known to weaken byssal threads by compromising the strength of the adhesive plaque. This study uses structural analysis to evaluate how an additional stressor, elevated temperature, influences byssal thread quality and production. Mussels (Mytilus trossulus) were placed in controlled temperature and pCO2 treatments, and then, newly produced threads were counted and pulled to failure to determine byssus strength. The effects of elevated temperature on mussel attachment were dramatic; mussels produced 60% weaker and 65% fewer threads at 25°C in comparison to 10°C. These effects combine to weaken overall attachment by 64-88% at 25°C. The magnitude of the effect of pCO2 on thread strength was substantially lower than that of temperature and, contrary to our expectations, positive at high pCO2 exposure. Failure mode analysis localized the effect of temperature to the proximal region of the thread, whereas pCO2 affected only the adhesive plaques. The two stressors therefore act independently, and because their respective target regions are interconnected (resisting tension in series), their combined effects on thread strength are exactly equal to the effect of the strongest stressor. Altogether, these results show that mussels, and the coastal communities they support, may be more vulnerable to the negative effects of ocean warming than ocean acidification.

Malpighian tubules of caterpillars: blending RNAseq and physiology to reveal regional functional diversity and novel epithelial ion transport control mechanisms.

The Malpighian tubules (MTs) and hindgut constitute the functional kidney of insects. MTs are outpouchings of the gut and in most insects demonstrate proximodistal heterogeneity in function. In most insects, such heterogeneity is confined to ion/fluid secretion in the distal portion and ion/fluid reabsorption in the proximal portion. In contrast, MTs of larval Lepidoptera (caterpillars of butterflies and moths) are composed of five regions that differ in their association with the gut, their structure and ion/fluid transport function. Recent studies have shown that several regions can rapidly and reversibly switch between ion secretion and reabsorption. The present study employed RNAseq, pharmacology and electrophysiology to characterize four distinct regions of the MT in larval Trichoplusia ni Luminal microelectrode measurements indicate changes in [K+], [Na+] and pH as fluid passes through different regions of the tubule. In addition, the regions examined differ in gene ontology enrichment, and demonstrate robust gradients in expression of ion transporters and endocrine ligand receptors. Lastly, the study provides evidence for direct involvement of voltage- and ligand-gated ion channels in epithelial ion transport of insect MTs.

Septate junction in the distal ileac plexus of larval lepidopteran Trichoplusia ni: alterations in paracellular permeability during ion transport reversal.

The Malpighian tubules (MTs) and hindgut together act as the functional kidney in insects. MTs of caterpillars are notably complex and consist of several regions that display prominent differences in ion transport. The distal ileac plexus (DIP) is a region of MT that is of particular interest because it switches from ion secretion to ion reabsorption in larvae fed on ion-rich diets. The pathways of solute transport in the DIP are not well understood, but one potential route is the paracellular pathway between epithelial cells. This pathway is regulated by the septate junctions (SJs) in invertebrates, and in this study, we found regional and cellular heterogeneity in the expression of several integral SJ proteins. DIP of larvae fed ion-rich diets demonstrated a reduction in paracellular permeability, coupled with alterations in both SJ morphology and the abundance of its molecular components. Similarly, treatment in vitro with helicokinin (HK), an antidiuretic hormone identified by previous studies, altered mRNA abundance of many SJ proteins and reduced paracellular permeability. HK was also shown to target a secondary cell-specific SJ protein, Tsp2A. Taken together, our data suggest that dietary ion loading, known to cause ion transport reversal in the DIP of larval Trichoplusiani, leads to alterations in paracellular permeability, SJ morphology and the abundance of its molecular components. The results suggest that HK is an important endocrine factor that co-regulates ion transport, water transport and paracellular permeability in MTs of larval lepidopterans. We propose that co-regulation of all three components of the MT function in larval lepidopterans allows for safe toggling between ion secretion and reabsorption in the DIP in response to variations in dietary ion availability.

secCl is a cys-loop ion channel necessary for the chloride conductance that mediates hormone-induced fluid secretion in Drosophila.

Organisms use circulating diuretic hormones to control water balance (osmolarity), thereby avoiding dehydration and managing excretion of waste products. The hormones act through G-protein-coupled receptors to activate second messenger systems that in turn control the permeability of secretory epithelia to ions like chloride. In insects, the chloride channel mediating the effects of diuretic hormones was unknown. Surprisingly, we find a pentameric, cys-loop chloride channel, a type of channel normally associated with neurotransmission, mediating hormone-induced transepithelial chloride conductance. This discovery is important because: 1) it describes an unexpected role for pentameric receptors in the membrane permeability of secretory epithelial cells, and 2) it suggests that neurotransmitter-gated ion channels may have evolved from channels involved in secretion.

Water and ion transport across the eversible vesicles in the collophore of the springtail Orchesella cincta.

Springtails (Collembola) are ancient close relatives of the insects. The eversible vesicles are their unique paired transporting organs, which consist of an epithelium located inside a tube-like structure called the collophore on the first abdominal segment. The vesicles can be protruded out of the collophore and several lines of evidence indicate that they have a vital function in water uptake and ion balance. However, the amount of water absorbed by the vesicles and which other ions apart from Na+ are transported remain unknown. Using Orchesella cincta as a model, we developed protocols for two assays that enabled us to study water and ion movement across the eversible vesicles in whole living springtails. Using an inverse Ramsay assay we demonstrate that the eversible vesicles absorb water from a droplet applied onto their surface. Using the scanning ion-selective electrode technique (SIET), we show that the vesicles absorb Na+ and Cl- from the bathing medium, secrete NH4+, and both absorb and secrete K+ H+ is secreted at a low level in the anterior part and absorbed at the posterior part. We did not detect transport of Ca2+ at significant levels. The highest flux was the absorption of Cl-, and the magnitude of ion fluxes was significantly lower in fully hydrated springtails. Our data demonstrate that the eversible vesicles are a transporting epithelium functioning in osmo- and ionoregulation, nitrogenous waste excretion and probably also acid-base balance.

Section-specific H+ flux in renal tubules of fasted and fed goldfish.

A recent study demonstrated that in response to a feeding-induced metabolic acidosis, goldfish (Carassius auratus) adjust epithelial protein and/or mRNA expression in their kidney tubules for multiple transporters known to be relevant for acid-base regulation. These include Na+/H+ exchanger (NHE), V-type H+-ATPase (V-ATPase), cytoplasmic carbonic anhydrase, HCO3- transporters and Rhesus proteins. Consequently, renal acid output in the form of protons and NH4+ increases. However, little is known about the mechanistic details of renal acid-base regulation in C. auratus and teleost fishes in general. The present study applied the scanning ion-selective electrode technique (SIET) to measure proton flux in proximal, distal and connecting tubules of goldfish. We detected increased H+ efflux into the extracellular fluid from the tubule in fed animals, resulting from paracellular back-flux of H+ through the tight junction. By applying inhibitors for selected acid-base regulatory epithelial transporters, we found that cytosolic carbonic anhydrase and HCO3- transporters were important in mediating H+ flux in all three tubule segments of fed goldfish. Contrastingly, V-ATPase seemed to play a role in H+ flux only in proximal and distal tubules, and NHE in proximal and connecting tubules. We developed working models for transport of acid-base relevant equivalents (H+, HCO3-, NH3/NH4+) for each tubule segment in C. auratus kidney. While the proximal tubule appears to play a major role in both H+ secretion and HCO3- reabsorption, the distal and connecting tubules seem to mainly serve for HCO3- reabsorption and NH3/NH4+ secretion.

Helicokinin alters ion transport in the secondary cell-containing region of the Malpighian tubule of the larval cabbage looper Trichoplusia ni.

Excretion in insects is accomplished by the combined actions of the Malpighian tubules (MTs) and hindgut, which together form the functional kidney. MTs of many insect groups consist of principal cells (PC) and secondary cells (SC). In most insect groups SCs are reported to secrete ions from haemolymph into the tubule lumen. Paradoxically, SCs in the MTs of the lepidopteran cabbage looper T. ni are used to reabsorb Na+ and K+ back into haemolymph. The current study was designed to investigate the effects and mode of action of the lepidopteran kinin, Helicokinin (HK), on ion transport in the SC-containing region of MT of T. ni. We identified a HK receptor (HK-R) homologue in T. ni and detected its expression in the SC-containing region of the MTs. The mRNA abundance of hk-r altered in response to changes in dietary K+ and Na+ content. HK-R immunolocalized to both PCs and SCs. Ramsay assays of preparations of the isolated distal ileac plexus (DIP) indicated that [HK] = 10-8 M: (i) decreased fluid secretion rate in unstimulated and serotonin-stimulated preparations, and (ii) increased [Na+]/[K+] ratio in the secreted fluid. Scanning ion-selective electrode technique measurements revealed that HK reduced: (i) K+ secretion by the PCs, and (ii) Na+ reabsorption by the SCs in intact tubules. In vitro incubation of the DIP with HK resulted in reduced mRNA abundance of hk-r as well as Na+/K+-ATPase subunit α (NKAα), Na+/K+/Cl- co-transporter (nkcc), Na+/H+ exchangers (nhe) 7 and 8, and aquaporin (aqp) 1. Taken together, results of the current study suggest that HK is capable of altering fluid secretion rate and [Na+]/[K+] ratio of the fluid, and that HK targets both PCs and SCs in the DIP of T. ni.