Physiology: Ion transport by human endometrial epithelia in vitro

Human glandular endometrial epithelial cells were cultured onporous tissue culture inserts to form tight, confluent layers.These layers generated time-dependent modifications in the ioniccomposition of both apical and basolateral solutions. Increasesin sodium and chloride concentrations in the basolateral fluidwere accompanied by reciprocal decreases in the concentrationsof these ions in the apical fluid. The potassium concentrationwas increased in the apical, while decreased in the basolateral,solution. The total calcium concentration was slightly elevatedin the apical, as compared with the basolateral fluid, whilethere were no alterations in pH. The endometrial layers demonstrateda significant transepithelial potential difference, and whenthis value was substituted in the Nernst equation a predictionof the passive distribution of ions across the cells was possible,indicating that none of the ions were in equilibrium. Additionof the sodium channel blocker amiloride to the medium bathingthe cell layers reduced the modifications in ionic compositionof apical and basolateral solutions. The data are consistentwith other data indicating an amiloride-sensitive sodiumabsorptivefunction for the endometrial epithelium. The ability of theseprimary cultures of endometrial epithelial cells to reduce thesodium while increasing the potassium concentration of the apicalfluid is qualitatively in agreement with the low sodium andhigh potassium concentrations reported for human uterine fluid.The data suggest a role for the endometrial epithelium in generatingand maintaining the distinctive ionic composition of the intra-uterineenvironment.     

Effects of nicotine on intestinal epithelial cells in vivo and in vitro: an X-ray microanalytical study

It has been suggested that nicotine may have a beneficial effect on ulcerative colitis. Little is known, however, about the mechanism of such an effect. In the present study, the effects of nicotine on intestinal ion transport were investigated by X-ray microanalysis. In addition to an in vivo study, an in vitro system was established by pre-incubating pieces of small intestine in solutions of different ionic composition, in an attempt to stabilize the intracellular ionic composition. It was observed that nicotine caused a loss of Cl (indicative of net chloride efflux) in the villous epithelial cells and the cells of Brunner's glands, in vitro both in Na+ and in K+-containing medium. In in vivo experiments, exposure of mice for 10 days to nicotine in drinking water resulted in a lowering of the cellular chloride concentration in villous epithelial cells of the small intestine, cells of Brunner's glands and epithelial cells of the colon. The results may indicate a disturbance of fluid transport in the intestine by nicotine. Possible mechanisms, such as inhibition of uptake of chloride across the basolateral membrane, are discussed.     

EBIO, an agent causing maintained epithelial chloride secretion by co-ordinate actions at both apical and basolateral membranes

The effect of 1-ethyl-2-benzimidazolone (EBIO) on electrogenic chloride secretion in murine colonic and nasal epithelium was investigated by the short-circuit technique. In the colon, EBIO produces a sustained current increase in the presence of amiloride, which is sensitive to furosemide. In nasal epithelium EBIO causes only a small, transient current increase. Sustained increases in current were obtained in response to forskolin in both epithelia. To examine the mechanisms by which EBIO increases chloride secretion, the effects on intracellular mediators were measured in colonic crypts. There was no effect on [Ca(2+)]i but cAMP content was increased, more so in the presence of IBMX, indicating a direct effect on adenylate cyclase. In colonic epithelia in which the apical surface was permeabilized by nystatin, and the tissue subjected to an apical to basolateral K(+) gradient, EBIO caused a current increase that was entirely sensitive to charybdotoxin (ChTX). In similarly permeabilized colons Br-cAMP caused a current increase that was entirely sensitive to 293B. Thus EBIO increases chloride secretion in the colon by coordinated actions at both the apical and basolateral faces of the cells. These include direct and indirect actions on Ca(2+)-sensitive and cAMP-sensitive K(+) channels respectively, and indirect actions on the basolateral cotransporter and apical CFTR chloride channels via cAMP. In CF colonic epithelia EBIO did not evoke chloride secretion. It is not clear why the nasal epithelium responds poorly to EBIO whereas it gives a sustained response to the related compound chlorzoxazone.     

Electrophysiology of cell volume regulation in proximal tubules of the mouse kidney

The present study has been designed to test for the influence of cell swelling on the potential difference and conductive properties of the basolateral cell membrane in isolated perfused proximal tubules. During control conditions the potential difference across the basolateral cell membrane (PD_bl) is –65±1 mV (n=74). Decrease of peritubular osmolarity by 80 mosmol/l depolarizes the basolateral cell membrane by +7.8±0.5 mV (n=42). An increase of bath potassium concentration from 5 to 20 mmol/l depolarizes the basolateral cell membrane by +25±1 mV (n=11), an increase of bath bicarbonate concentration from 20 to 60 mmol/l hyperpolarizes the basolateral cell membrane by –3.2±0.5 mV (n=13). A decrease of bath chloride concentration from 79.6 to 27 mmol/l hyperpolarizes the basolateral cell membrane by –1.8±0.7 mV (n=6). During reduced bath osmolarity, the influence of altered bath potassium concentration on PD_bl is decreased ( PD_bl=+16±2 mV,n=11), the influence of altered bicarbonate concentration on PDbl is increased ( PD_bl=–6.0±0.8 mV,n=13), and the influence of altered bath chloride concentration on PD_bl is unaffected ( PD_bl=–1.8±0.6 mV,n=6). Barium depolarizes the basolateral cell membrane to –28±2 mV (n=16). In the presence of 1 mmol/l barium, decrease of peritubular osmolarity by 80 mosmol/l leads to a transient hyperpolarization of the basolateral cell membrane by –5.9±0.5 mV (n=16). This transient hyperpolarization is blunted in the absence of extracellular bicarbonate. In conclusion, cell swelling depolarizes straight proximal tubule cells and increases bicarbonate selectivity of the basolateral cell membrane at the expense of potassium selectivity. The data reflect either incrases of bicarbonate conductance or decrease of potassium conductance during exposure of proximal tubule cells to hypotonic media.Parts of this work were presented at the 18th Congress of the Gesellschaft für Nephrologie, Frankfurt/M. 1986 and at the 8th International Symposium on Biochemical Aspects of Kidney Function, Dubrovnik 1986     

1-EBIO stimulates Cl– secretion by activating a basolateral K+ channel in the mouse jejunum

We investigated the effects of 1-ethyl-2-benzimidazolinone (1-EBIO) on ion transport in the mouse jejunum through the use of the short-circuit (Isc) current technique and the application of the patch-clamp technique to isolated jejunal crypts. In HCO3- Ringer's, 1-EBIO stimulated a dose-dependent (EC50 964 micromol/l), bumetanide-sensitive increase in Isc consistent with stimulation of Cl- secretion. In contrast, in Cl(-)-free HCO3-Ringer's containing glucose, 1-EBIO (500 micromol/l) did not increase the phloridzin (100 micromol/l) sensitive Isc, suggesting that electrogenic Na+ absorption was unaltered. Measurement of the membrane potential (Vm) with the perforated-patch technique indicated that in isolated crypts, 1-EBIO caused a reversible hyperpolarization of Vm and an increase in the change in Vm associated with step changes in bath K+, consistent with an increase in K+ conductance. In on-cell patch experiments with KCI Ringer's in the patch pipette and crypts bathed with NaCl Ringer's, 1-EBIO (500 micromol/l) increased the open probability (NPo; 0.01+/-0.01 to 0.45+/-0.11, n=7) of an inwardly rectified intermediate conductance (g) channel. In inside-out patches with KCl Ringer's in the patch pipette and KCI Ringer's containing 100 nmol/l Ca2+ in the bath, the current-voltage relationship of the channel was inwardly rectified (g of 10 and 52 pS at -Vp of 100 and -100 mV, respectively) and reversed at 0 mV (n=5). Replacement of bath K+ with Na+ shifted the reversal potential toward the equilibrium potential for K+. In the presence of 1-EBIO, reducing the bath Ca2+ from 200 nmol/l to nominally Ca(2+)-free conditions decreased NPo from 0.90+/-0.27 to 0.07+/-0.03 (n=3). We conclude that in the mouse jejunum, I-EBIO does not stimulate electrogenic Na+ absorption. It does, however, stimulate secretion primarily through the activation of a basolateral, intermediate conductance Ca(2+)-sensitive K+ channel.     

Chloride transport by self-exchange and by KCI salt diffusion in gramicidin-treated human red blood cells

The permeability of gramicidin-treated human red blood cell membranes to K⁺ and CI^- has been measured at normal ionic strength (1) by tracer exchange at steady-state distribution of salt, and (2) by net transport of salt in the presence of a salt concentration gradient. Under both conditions KCI was the only inorganic salt in cells and medium. In the studies of self-exchanges the electrical driving force on the ions was zero. Calculation of permeability coefficients from net salt transport was simplified because the experiment was designed as a special case of the Nernst-Planck diffusion regime, i.e. the single salt case. Gramicidin altered the cell membranes from being anion to become cation selective. Gramicidin increased the potassium exchange without affecting, the chloride exchange measurably. The chloride exchange showed saturation kinetics as does chloride exchange in normal cells. The net transport of KCI in the presence of a constant concentration gradient increased to a constant value with increasing gramicidin concentration. At high gramicidin concentrations (0°C, pH 7.2) the “chloride permeability coefficient” calculated from tracer exchange (1.9×10^-6 cm/s) was 290 times the chloride permeability coefficient calculated from net salt transport (0.65×10^-8 cm/s). The latter value corresponds to a chloride conductance of 4.2×10^-6 ohm^-1 cm^-2. The chloride permeability coefficient was 2.1×10^-8 cm/s at 25°C (pH 6.8) indicating a value of 3 for the Q₂₅. It appears that normal red cells are anion selective in the sense that anion permeability exceeds cation permeability with a factor of more than a hundred between 0°C and body temperature. The anion exchange, i.e. the Hamburger shift, is a tightly coupled transport process which is several orders of magnitude faster than anion transport by salt diffusion.     

Evidence for basolateral membrane potassium conductance in canine tracheal epithelium

The ionic dependence of the basolateral membrane conductance in canine tracheal epithelium was investigated using intracellular microelectrode techniques. Increasing the K+ concentration in the submucosal bathing solution depolarized the electrical potential difference across the basolateral membrane; neither alteration of the submucosal Na+ concentration nor the mucosal K+ concentration had a significant effect on the cellular electrical potential profile. An increase in the K+ concentration in the submucosal bathing solution also decreased the net rate of Cl-secretion. Addition of ouabain (10(-4) M) to the submucosal bathing solution decreased the short-circuit current and depolarized the intracellular voltage without altering transepithelial resistance or the cell membrane resistance ratio, suggesting that basolateral resistance was unchanged. These findings, together with the previous observation that there is no appreciable basolateral Cl- conductance, indicate that a K+ conductance accounts for the predominance of the electrical conductance at the basolateral membrane. The results also indicate that the basolateral membrane K+ conductance plays a critical role in the generation of the negative intracellular voltage that drives Cl- exit across the apical membrane and thus supports Cl- secretion.     

Ion-secreting epithelia: chloride cells in the head region of Fundulus heteroclitus

Transporting cells of ion-secreting epithelia are characterized by similar morphological patterns that include rich supplies of mitochondria, exotic patterns of surface amplification, and basolateral, plasma-membrane location of Na-K-ATPase, even though the direction of sodium transport across these epithelia is toward the apical side. Several new models for NaCl secretion propose that sodium, extruded into the intercellular space by Na-K-ATPase, reaches the apical side via the zonulae occludentes. Very recent freeze-fracture electron microscopy of avian salt gland and teleost chloride cells reveals that transporting cells are joined by simple, shallow zonulae occludentes. These observations lend morphological support to the concept that paracellular sodium ion permeation plays a central role in secretion. The chloride ion may traverse the epithelium via a transcellular route, entering the cell at the basolateral membrane by a chloride carrier linked to the cotransport of sodium down its electrochemical gradient into the cell. Finally, the chloride ion may exit the cell across the apical membrane by electrical forces. This review summarizes biochemical, morphological, and electrophysiological aspects of these new secretory models and the important contribution of a half century of research on teleost osmoregulatory mechanisms, including the chloride cell, to our understanding of sodium and chloride transport across secretory epithelia.     

Role of Na-K-ATPase in chloride cell function

In seawater eels the efflux of sodium (and chloride) across the gill is directly proportional to the activity of Na-K-ATPase in homogenates of gill filaments. The rate of ion movement, however, is substantially greater than at the temperature of seawater. Na-K-ATPase is localized predominantly on the basolateral surface of the chloride cell so that ouabain inhibits from the blood side rather than from the apical or mucosal surface of chloride cells. Chloride, rather than sodium, is probably the actively transported ion species, and an attractive hypothesis for active chloride transport is one that invokes the cotransport of chloride with sodium across the basolateral membrane, the energy for which is supplied indirectly by the operation of the Na-K-ATPase pump. Exposure to freshwater sharply dissociates ion movements from Na-K-ATPase activity, possibly by changing the permeability of cell membranes to chloride movements.     

Ultrastructural features of chloride cells in the gill epithelium of the atlantic salmon,Salmo salar,and their modifications during smoltification

To elucidate the ultrastructural modifications of the gill epithelium during smoltification, gills of the Atlantic salmon (Salmo solar) were examined by electron microscopy at three stages of this process, which were defined as follows: “parrs” were freshwater fish that had not yet started their transformation; “freshwater smolts” were freshwater fish that were ready to enter seawater; and “seawater smolts” were smolts that had been transferred from fresh water and maintained for 4 days in seawater (35%). In the gill epithelium of parrs, there were two types of chloride cells. The large chloride cells contained deeply stained mitochondria and numerous apical, irregular, dense, membrane-bound bodies that formed 77% of the chloride cell population and were distinguished easily from small chloride cells that have distinctly paler mitochondria and no dense bodies in their apical cytoplasm. In freshwater smolts, the large chloride cells formed 95% of the chloride-cell population. In contrast to the small chloride cells that were not modified, they almost doubled in size. Their tubular system developed extensively to form a tight network with regular meshes significantly smaller than those observed in parr chloride cells. Forty percent of the large chloride cells were associated with a new type of cell, the accessory cell, to which they were bound by shallow apical junctions. Half of these accessory cells were not seen to be in contact with the external medium, In seawater smolts, 80% of the large chloride cells were associated with accessory cells. Most accessory cells reached the external medium and sent numerous cytoplasmic interdigitations within the apical portion of the adjacent chloride cells. As a result, a section through the apical portion of the chloride cells and their associated accessory cells revealed a mosaic of interlocked cell processes bound together by an extended, shallow apical junction. It was concluded that the Atlantic salmon develops in fresh water most of the ultrastructural modifications of the gill epithelium which in most euryhaline fish are triggered by exposure to seawater. The effective transfer into seawater would act only as a final stimulus to achieve some adequacy between the freshwater smolt and its new environment.     

Current mechanisms of macula densa cell signalling

Macula densa cells couple renal haemodynamics, glomerular filtration and renin release with tubular fluid salt and water reabsorption. These cells detect changes in tubular fluid composition through a complex of intracellular signalling events that are mediated by membrane transport pathways. Increases in luminal fluid sodium chloride concentration result in alterations in cell sodium chloride concentration, cytosolic calcium, cell pH, basolateral membrane depolarization and cell volume. Macula densa signalling then involves the production and release of specific paracrine signalling molecules at their basolateral membrane. Upon moderate increases in luminal sodium chloride concentration macula densa cells release increasing amounts of ATP and decreasing amounts of prostaglandin E(2), thereby increasing afferent arteriolar tone and decreasing the release of renin from granular cells. On the other hand, further increases in luminal concentration stimulate the release of nitric oxide, which serve to prevent excessive tubuloglomerular feedback vasoconstriction. Paracrine signalling by the macula densa cells therefore controls juxtaglomerular function, renal vascular resistance and participates in the regulation of renin release.     

Intracellular potassium activity in the rabbit proximal straight tubule

Double-barreled liquid ion-exchanger microelectrodes were used to measure basolateral membrane potential (VBL) and intracellular potassium activity (aiK) in superficial proximal straight tubules (sPST) of the rabbit perfused in vitro. The mean +/- SE (number of cells in parentheses) value of VBL was -37.8 +/- 2.49 (20) vM and aiK was 48.6 +/- 2.27 (20) mM. The calculated Nernst equilibrium potential (EK) across the basolateral membrane was -68 mV. Lowering both potassium concentration to 0.1 mM reversibly decreased both VBL and aiK to -12.2 +/- 1.21 (19) mV and 11.3 +/- 1.29 (19) mM, respectively. Bath ouabain (10(-5) resulted in similar changes. These results demonstrate that intracellular potassium is actively accumulated in sPST perfused in vitro and that accumulation results primarily from Na-K-ATPase activity in the basolateral membrane. During recovery from low K bath, the temporal relationship between VBL and aiK and the effects of ouabain and high K bath on recovery are used to demonstrate directly electrogenic pumping. Lowering bath pH to 6.7 (HCO-3 = 5 mM) and the presence of 0.5 mM BaCl2 in the bath resulted in a large and rapid depolarization of VBL with little or no change in aiK. These results suggest that the response of VBL to both maneuvers is caused by a decrease in potassium permeability of the basolateral membrane.     

Ultrastructural features of mitochondria-rich cells in stenohaline freshwater and seawater fishes

In order to elucidate the functional significance of accessory cells in freshwater fishes, such as the rainbow trout, which displays a poor adaptability to seawater life, a search for such cells was performed in two stenohaline freshwater fishes: the loach and the gudgeon. Accessory cells were never encountered in these species; but, in contrast, two types of chloride cells were observed consistently that strikingly resembled the α- and β-cells previously described in the guppy, a freshwater-adapted euryhaline fish. The α-cell, a pale and elongated chloride cell, was located at the base of the secondary lamellae in close contact with the arterioarterial pillar capillary. Darker, ovoid chloride cells resembling the β-cell were found exclusively in the interlamellar region of the primary epithelium facing the central venous sinous. The latter cells frequently formed multicellular complexes linked together by deep, narrow, apical junctions. In another experiment, a stenohaline seawater fish, the turbot, was adapted to diluted 5% saltwater and to fresh water. In seawater, the gill epithelium contained only one type of chloride cell, always associated with accessory cells. Due to numerous cytoplasmic interdigitations between the accessory cells and the apical portion of the chloride cell, there was a noticeable increase in the length of the shallow apical junction, sealing off the intercellular space between the two cell types. In 5% saltwater, there was a decrease in the number of these interdigitations and a concomitant decrease in the length of the shallow apical junction. In fresh water, chloride cells were partially or completely separated from the outside medium by modified accessory cells. It is thus concluded that accessory cells are found exclusively in fish living in seawater or preadapted to seawater and that they probably are involved in the formation and modulation of paracellular pathways for ionic excretion. In contrast, the respective roles of the two types of chloride cells observed in freshwater fishes are still to be determined.     

Intracellular pH in isolated rat renal papillary thin limbs of Henle's loop

Intracellular pH (pHi) was measured in isolated, nonperfused and perfused rat papillary thin limbs of Henle's loops in N-2-hydroxyethylpiperazine-N'-2-ethansulfonic acid (HEPES)- or HEPES/bicarbonate-buffered medium at pH 7.4 using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Resting pHi was about 6.7 in descending thin limbs (DTL) and about 6.9 in ascending thin limbs (ATL), even with a medium pH of 7.4. These values appeared to reflect the acid pH of the blood in the neighboring vasa recta found in vivo. The resting pHi did not differ whether or not the medium contained bicarbonate although the total buffering capacity of the tubule cells was increased in the presence of bicarbonate. In nonperfused DTL and ATL, pHi was further acidified following an NH4Cl pulse. The rate of recovery of pHi from this level to the resting pHi was reduced by Na+ removal from the bath in both DTL and ATL and by the addition of ethylisopropylamiloride (EIPA) to the bath in the presence of Na+ in DTL. The rate of recovery was not affected by Cl- removal from the bath or K+ (75 mM) or 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) addition to the bath in either DTL or ATL. These results suggest that the common, amiloride-sensitive, basolateral Na+/H+ exchanger plays a role in the regulation of pHi in rat papillary DTL but that a different basolateral Na+/H+ exchanger or a luminal Na+/H+ exchanger is important in rat papillary ATL.     

Cell volume-induced changes in K+ transport across the rat colon.

The effect of cell swelling and cell shrinkage on K+ transport across the rat colonic epithelium was studied by measuring unidirectional fluxes, uptake and efflux of 86Rb+, a marker for K+. Exposure to a hypotonic medium stimulated the secretory, serosa-to-mucosa flux of K+, whereas exposure to a hypertonic medium inhibited the absorptive, mucosa-to-serosa flux of K+ in the distal, but not in the proximal colon. Neither manoeuvre had any effect on the uptake of K+ across the apical or the basolateral membrane. Cell swelling induced a sustained increase in the apical and basolateral K+ efflux from both colonic segments, whereas cell shrinkage reduced the efflux. Ba2+ (10(-2) mol l(-1)) inhibited the swelling-induced stimulation of the apical, quinine (10(-3) mol l(-1)) that of the basolateral K+ efflux in the distal colon. Incubation of the tissue in Ca2+-free buffer or La3+, which blocks Ca2+-influx into the epithelium, strongly reduced the basal K+ efflux across the basolateral membrane. The same was observed with brefeldin A, a blocker of the transport of newly synthesized proteins out of the endoplasmatic reticulum. Swelling-induced K+ efflux, however, was not reduced. In the presence of colchicine, an inhibitor of the polymerization of microtubules, swelling evoked only a transient increase in mucosal efflux, which, especially in the proximal colon, fell after 6 min to the level of the isotonic control period. These results demonstrate that the cell volume is involved in the regulation of transepithelial K+ transport across the rat colonic epithelium and suggest a role of the cytoskeleton in the control of a part of the volume-sensitive K+ channels.     

Functional heterogeneity in the hamster medullary thick ascending limb of Henle's loop

Cellular heterogeneity was examined in the hamster medullary thick ascending limb (MAL) perfused in vitro by electrophysiological measurements with an intracellular microelectrode. Random measurements of fractional resistance of basolateral membrane (Rf _B) revealed two cell populations, high basolateral conductance (HBC) cells havingRf _B of 0.05±0.01 (n=24) and low basolateral conductance (LBC) cells havingRf _B of 0.80±0.03 (n=32). Basolateral membrane potentials (V _B) were not different between HBC cells and LBC cells (–72.6±1.2,n=43 vs. –70.0±1.2,n=35). Addition of 2 mmol/l Ba²⁺ to the bath depolarized the basolateral membrane in the HBC cells from –70.4±3.2 to –20.9±5.9 mV (n=8) but not in the LBC cells (from –74.4±1.9 to –72.0±2.1 mV). Increasing K⁺ or decreasing Cl^– in the bathing solution caused marked positive deflection ofV _B in the HBC cells but little or no change inV _B in the LBC cells. Elimination of Cl^– from the lumen or addition of furosemide to the lumen enhanced the potential response of the HBC cells to basolateral application of Ba²⁺. Accordingly, with Ba²⁺ present in the bath, the potential response of the HBC cells to a decrease in bath Cl^– concentration was enhanced. These observations suggest that a K⁺ conductance exists in the basolateral membrane of HBC cells in paralled with a Cl^– conductance. The basolateral cell membrane of LBC cells also contains a Cl^– conductance. In these cells, but not in HBC cells, the potential response to decreasing bath Cl^– concentration increased when bath pH was decreased from 7.4 to 6.0 Apparent K⁺ transference numbers of the luminal membrane were higher in LBC cells (0.74±0.05,n=7) than in HBC cells (0.20±0.02,n=5). From these data, we conclude: (1) there are two distinct cell types in the hamster medullary thick ascending limb; (2) there is a low Cl^– conductance in basolateral membrane of LBC cells which is stimulated by low pH.     

Immunohistochemical localization of sodium-potassium-stimulated adenosine triphosphatase and carbonic anhydrase in human colon and colonic neoplasms.

Sodium-potassium-stimulated adenosine triphosphatase and carbonic anhydrase isozymes I and II were localized immunocytochemically in adenomas, adenocarcinomas, and normal epithelium of human colon harboring non-neoplastic lesions. Non-neoplastic control colon showed carbonic anhydrase I and II in the cytoplasm of the columnar cells lining the upper half of the crypts. Antiserum to sodium-potassium-stimulated adenosine triphosphatase bound to the basolateral but not the apical plasmalemma of columnar epithelial cells. Staining was most intense in the superficial cells, which also contained carbonic anhydrase, but was also evident to a lesser degree in cells deep in the crypts. Adenomas and adenocarcinomas failed to stain for content of carbonic anhydrase but retained basolateral sodium-potassium adenosine triphosphatase positivity. The staining characteristics of colonic neoplasms for the two enzymes involved in the transport function of colonic epithelium thus resembled those of the less mature cells lining the base of normal crypts.     

Evidence from O2 uptake measurements for Na+−K+−2 Cl− co-transport in the rabbit submandibular gland

There is evidence that the production of primary saliva by acinar cells is a consequence of Na⁺–Cl^– co-transport but more recently it has been proposed that in fact Na⁺–K⁺–2 Cl^– co-transport is responsible. The latter would be energetically more efficient and the present experiments were designed to measure the stoichiometry of acinar secretion in order to distinguish between these two mechanisms.Submandibular salivary glands from anaesthetised rabbits were isolated vascularly and oxygen consumption measured from the oxygen content of arterial inflow and venous effluent blood and the total flow through the gland. Measurements were made in the steady-state at rest and during different secretion rates induced by parasympathetic nerve stimulation. The rate of sodium transport across the acinar and ductal epithelium was determined from plasma and salivary sodium concentration and salivary flow rate.Multiple regression analysis of this data showed that 22.1 mol Na⁺ was secreted per mol O₂ consumed while 11.9 mol Na⁺ was reabsorbed per mol O₂ consumed. Since acinar secretion is energetically about twice as efficient as ductal absorption, a mechanism for Na⁺ transport other than that for tight epithelia must be involved. Na⁺K⁺–2 Cl^– co-transport is thus more likely than Na⁺–Cl^– and it is suggested that Na⁺–K⁺–2 Cl^– co-transport is the main mechanism involved in salivary acinar secretion.     

Sodium, bicarbonate, and chloride absorption by the proximal tubule

Proximal tubules are lined with epithelial cells that contain Na-K-ATPase in their basolateral cell membrane. The luminal cell membrane contains transport proteins that couple movement of many solutes to the active transport of sodium. The cells are connected by low-resistance junctional complexes that permit passive movement of solutes via a paracellular shunt pathway. Acidification is mediated by a Na/H antiporter localized specifically in the luminal membrane and a chloride-independent, voltage-dependent bicarbonate exit process in the basolateral membrane. The rate of acidification is controlled by the pH of the luminal and peritubular fluids. Reabsorption of NaCl from the high-chloride, low-bicarbonate fluid in the late proximal tubule is approximately 40% passive and 60% active. In proximal straight tubules the active component is entirely by simple rheogenic sodium transport, with chloride absorption driven through the paracellular shunt pathway by the lumen-negative PD. In convoluted tubules the active component is primarily neutral, with both sodium and chloride transported in approximately equivalent amounts through the cell. The mechanisms for neutral NaCl transport across the luminal membrane and for chloride exit across the basolateral membrane are unknown. A reduction in peritubular Starling forces (hydraulic and oncotic pressures) suppresses net proximal reabsorption by two mechanisms: 1) increased paracellular permeability with modest backleak of solutes (bicarbonate, glucose, amino acids) whose luminal concentration falls below their plasma level, and 2) specific inhibition of active neutral transcellular transport of NaCl by reduced peritubular protein concentration by some mechanism other than inhibition of Na-K-ATPase.