Lateral Intercellular Spaces Research Articles

A stereological analysis of the effect of buffer osmolarity on a salt secreting epithelium, under variable physiological conditions

SUMMARYA stereological analysis of salt gland epithelium showed differences in the size of the intracellular space and the volume fraction of some cellular organelles, according to the salinity in which the animals were kept. These changes were similar to earlier non‐quantitative work on several salt secreting epithelia in various states of activity.However, alteration of one variable in the processing technique, osmolarity of the fixative buffer, was shown to produce the same stereological result in salt glands held at constant activity. Most experimental manipulations used in previous experiments to alter glandular activity, e.g. salt loading, altered perfusate, or inclusion of pump inhibitors, probably also alter intracellular ion contents. It is therefore difficult to decide whether changes in lateral intercellular space geometry represent physiological mechanisms in vivo or differences in osmotic behaviour during processing for electron microscopy.

Membrane differentiation in the cytoplasmic-tubule system of the intestinal absorptive cells of the lamprey, Lampetra japonica

Specific membrane differentiation occurs in the cytoplasmic-tubule system of the absorptive cells lining the mucosa of the lamprey anterior intestine. The absorptive cells are characterized by the presence of abundant mitochondria and a system of well-developed cytoplasmic tubules (∼120 nm in diameter). The cytoplasmic tubules open on to the basolateral cell surface and contain numerous lipoprotein particles (50–100 nm diam.) in their lumina. Lipoprotein particles are also observed in the endoplasmic reticulum and the Golgi complex, and they are transfered to the lateral intercellular space and lamina propria by way of the cytoplasmic tubules. Spirally-wound parallel rows of particles are found in the luminal surface of the cytoplasmic tubules. The rows are ∼17 nm apart and are wound spirally at a pitch of ∼210 nm. Freeze-fracture images of the tubule membranes also show spiral arrays of particles (∼9 nm in diameter) on the P-face, and complementary shallow grooves on the E-face. From these observations, it is suggested that the cytoplasmic-tubule system of the intestinal absorptive cells serves as a channel for the transport of synthesized lipoprotein into the interstitium, and is also the site of the ion and water exchange essential for the maintenance of ionic homeostasis.

Effects of colchicine on the gallbladder of the mouse.

The effects of colchicine on the mouse gallbladder followed a course depending on the dosage given (0.4-4 mg/100 g body weight). Following 0.5 mg/100 g, by 16 h there was a marked cholestasis with dilatation of the gallbladder and steatosis. There were progressive alterations in the Golgi apparatus and accumulation of vesicles. The apical mucous droplets decreased in number and became pleomorphic and dispersed throughout the cytoplasm. Lipid droplets appeared in numbers on the epithelial cytoplasm. By 48 h the tissues had reverted to normal appearances. When cholecystokinin, pilocarpine or ceruletide were given to animals which had received colchicine 18 h previously, the excess bile from the dilated gallbladder was discharged into the duodenum, remaining apical mucous droplets secreted and electron dense material accumulated in the lateral intercellular space. This formed a quasi-regular array between the epithelial bases and the basement membrane. Biochemically there was a significant decrease in alkaline phosphatase activity and a significant increase in acid phosphatase activity.

The roles of paracellular and transcellular pathways and submucosal space in isotonic water absorption by rabbit ileum.

Water movements have been studied in sheets of isolated rabbit ileum using a method which measures net volume flows across the mucosal and serosal surfaces of the tissue continuously with high resolution. At 35 degrees C, with the tissues incubated in isotonic Ringer solution containing D-glucose (25 mM) on both sides, there is a steady net inflow of fluid at the rate of 24 +/- 2 microliter cm-2 h-1 across the mucosal surface (Jm) and an outflow of 8 +/- 1 microliter cm-2 h-1 across the serosal surface (Js) (n = 16). The stable transepithelial p.d. across these tissues is 2.7 +/- 0.2 mV, serosa positive. Jm can be reversibly inhibited by anoxia. Ouabain (0.1 mM) added to the serosal solution inhibits inflow across the mucosal and serosal surfaces by 75% (n = 7) within 30 min. If phlorizin (0.1 mM) is added to the mucosal Ringer solution containing glucose (20 mM) within 30 min of the commencement of in vitro absorption, Jm is reduced from 37 +/- 3 to 28 +/- 2 microliter cm-2 h-1 (n = 3). Dilution of the mucosal Ringer solution by 50 mosmol kg-1 (with the serosal solution kept isosmolar) results in a rapid transient increase in mucosal inflow. An increase of 50 mosmol kg-1 in the mucosal Ringer solution with NaCl, sucrose or mannitol causes a transient reversal of mucosal flow, followed by a return of inflow at a reduced level. Rabbit ileum can transport water against gradients of approximately 75 mosmol kg-1 of sucrose, NaCl, or mannitol. Addition of polyethylene glycol (mol. wt. 20000; 3 mosmol kg-1) causes a sustained reversal of mucosal inflow; inflow can be restored only by removing polyethylene glycol from the mucosal Ringer solution. The tissue can absorb water against an osmotic gradient of 200 mM-glycerol. The above data have been incorporated into a new model to explain isotonic flow of fluid by this epithelium. The main features are that the hydraulic conductivity (Lp) of the mucosal boundary of the lateral intercellular space is approximately 1 X 10(-8) cm s-1 cmH2O-1. This Lp is too low to sustain isotonicity of the flow emerging from the lateral intercellular space at the observed rates. Hypertonic fluid emerging from the lateral intercellular space is diluted by transcellular water flow generated by the hypertonicity of the submucosa and back-diffusion of solute via mucosal shunt channels.(ABSTRACT TRUNCATED AT 400 WORDS)

Structural changes associated with fluid absorption by dog tracheal epithelium.

During fluid absorption induced by amphotericin B, the lateral intercellular spaces (LIS) of the dog tracheal epithelium were widely dilated as compared to untreated time controls. When fluid absorption was inhibited by ouabain, or by replacement of luminal Na by choline, amphotericin B failed to cause dilation of the LIS. These data suggest that, as in other epithelia, a significant amount of transepithelial fluid flow passes down the LIS, and that these spaces may provide the local osmotic compartment which is responsible for linking transepithelial fluid movement to active ion transport.

Antidiuretic hormone response in the amphibian urinary bladder: Time course of cytochalasin-induced vacuole formation, an ultrastructural study employing ruthenium red

Cytochalasin is known to inhibit the antidiuretic hormone-induced hydro-osmotic response (bulk water flow) in the amphibian urinary bladder without altering hormone-stimulated diffusional water permeability or short-circuit current. In addition, histological studies have shown that the mold metabolite induces the formation of large intracellular vacuoles or lakes in the epithelial cells. We report here a transmission electron microscopic time-course study which indicates that during the early phases of the ADH response cytochalasin causes the formation of numerous multivesicular bodies or aggregates derived from individual basolateral pinocytotic vesicles. Because of their apparent hypertonic nature, the vesicles, as well as the vesicular aggregates, accumulate water during hormone-stimulated hydro-osmotic flow. As a result, the multivesicular bodies dilate and fuse to form the large intracellular lakes characteristic of cytochalasin treatment in the presence of both an applied osmotic gradient and vasopressin. In the presence of mucosal ruthenium red, the luminal glycocalyx was heavily stained with this tracer. At no time, however, even in the presence of hormone, was there any evidence for the uptake of this dye at the apical epithelial border. In the presence of serosal ruthenium red, the lateral intercellular spaces, basolateral pinocytotic vesicles, basal lamina, and collagen, as well as other subepithelial structures, were ruthenium positive. With cytochalasin D, vasopressin, and serosal ruthenium red, both the pinocytotic vesicles and the multivesicular bodies demonstrated an apparent membrane associated ruthenium positive coat. The tracer data indicates that the basolateral pinocytotic vesicles, increased by the presence of hormone, are indeed endocytotic in nature. The mucopolysaccharide coat associated with these structures may be involved in ionic and/or fluid transport.

Response of the tracheobronchial epithelium to hemoprotein tracers.

The response of the tracheobronchial epithelium to intratracheal administration of microperoxidase, cytochrome c, myoglobin, and horseradish peroxidase (HRP) was investigated in hamsters and gerbils. Using diaminobenzidine cytochemistry and transmission electron microscopy, we found that cytochrome c was the only tracer to reach lateral intercellular spaces, most probably due to its toxic effect on the airway epithelium. Myoglobin did not bind to the plasma membrane and hence was not internalized by the epithelial cells. Both HRP and microperoxidase were seen in contact with the plasma membrane and within pinocytotic vesicles, the fate of which was not apparent. As early as 5 min after administration, HRP was distributed in a diffuse and intense manner in a small percentage of ciliated and nonciliated cells but by 30 min this effect had decreased significantly. HRP was seen in the basement membrane and lamina propria beneath some intensely reactive cells at 5 and 30 min. Whether these cells reflect a degenerate condition and hence are incapable of excluding HRP or whether HRP is itself toxic is unknown. The rapid rate at which these cells appear raises the possibility that they may represent a conduit that allows small but significant amounts of foreign molecules to reach subepithelial mast cells thus triggering or augmenting the Type I hypersensitivity response.

Implications of an anomalous intracellular electrical response in bullfrog corneal epithelium.

The ionic dependencies of the transepithelial and intracellular electrical parameters were measured in the isolated frog cornea. In NaCl Ringer's the intracellular potential difference Vsc measured under short-circuit conditions depolarized by nearly the same amount after either increasing the stromal-side KCl concentration from 2.5 to 25 mM or exposure to 2 mM BaCl2 (K+ channel blocker). With Ba2+ the depolarization of the Vsc by 25 mM K+ was reduced to one-quarter of the control change. If the Cl-permselective apical membrane resistance Ro remained unchanged, the relative basolateral membrane resistance Ri, which includes the lateral intercellular space, increased at the most by less than twofold after Ba2+. These effects in conjunction with the depolarization of the Vsc by 62 mV after increasing the stromal-side K+ from 2.5 to 100 mM in Cl-free Ringer's as well as the increase of the apparent ratio of membrane resistances (a = Ro/Ri) from 13 to 32 are all indicative of an appreciable basolateral membrane K+ conductance. This ratio decreased significantly after exposure to either 25 mM K+ or Ba2+. The decline of Ro/Ri with 25 mM K+ appears to be anomalous since this decrease is not consistent with just an increase of basolateral membrane conductance by 25 mM K+, but rather perhaps a larger decrease of Ro than Ri. Also an increase of lateral space resistance may offset the effect of decreasing Ri with 25 mM K+. In contrast, Ro/Ri did transiently increase during voltage clamping of the apical membrane potential difference Vo and exposure to 25 mM K+ on the stromal side.(ABSTRACT TRUNCATED AT 250 WORDS)

Epithelial water transport in a balanced gradient system

The relationship between epithelial fluid transport, standing osmotic gradients, and standing hydrostatic pressure gradients has been investigated using a perturbation expansion of the governing equations. The assumptions used in the expansion are: (a) the volume of lateral intercellular space per unit volume of epithelium is small; (b) the membrane osmotic permeability is much larger than the solute permeability. We find that the rate of fluid reabsorption is set by the rate of active solute transport across lateral membranes. The fluid that crosses the lateral membranes and enters the intercellular cleft is driven longitudinally by small gradients in hydrostatic pressure. The small hydrostatic pressure in the intercellular space is capable of causing significant transmembrane fluid movement, however, the transmembrane effect is countered by the presence of a small standing osmotic gradient. Longitudinal hydrostatic and osmotic gradients balance such that their combined effect on transmembrane fluid flow is zero, whereas longitudinal flow is driven by the hydrostatic gradient. Because of this balance, standing gradients within intercellular clefts are effectively uncoupled from the rate of fluid reabsorption, which is driven by small, localized osmotic gradients within the cells. Water enters the cells across apical membranes and leaves across the lateral intercellular membranes. Fluid that enters the intercellular clefts can, in principle, exit either the basal end or be secreted from the apical end through tight junctions. Fluid flow through tight junctions is shown to depend on a dimensionless parameter, which scales the resistance to solute flow of the entire cleft relative to that of the junction. Estimates of the value of this parameter suggest that an electrically leaky epithelium may be effectively a tight epithelium in regard to fluid flow.

Direct visualization of the isolated and perfused macula densa.

Direct examination of the structure and function of the macula densa is compromised by the relative inaccessibility and small size of this cell plaque. We report the isolation and perfusion of rabbit nephron segments with attached glomeruli and the direct visualization of the macula densa with differential interference-contrast microscopy. We used this technique to examine the structural sensitivity of the macula densa to perturbations in luminal osmolality and NaCl concentration. Reducing luminal osmolality from 290 to 70 mosmol/kg by removing NaCl resulted in a dilation of the lateral intercellular spaces that was both reversible and specific to the region of the macula densa. Associated with the dilation of the intercellular spaces was a small (congruent to 10%), but statistically significant, increase in the height of the macula densa cells. These structural events were related to the reduction in luminal osmolality, since isosmotic replacement of NaCl with mannitol resulted in no detectable structural changes. Thus, the macula densa may represent a small water-permeable plaque of cells within the remaining water-impermeable thick ascending limb of Henle's loop.

Ion transport by primary cultures of canine tracheal epithelium: methodology, morphology, and electrophysiology.

Canine tracheal epithelial cells were isolated by enzymatic and mechanical dispersion and cultured on permeable supports. The cells formed confluent monolayers and retained most of the morphologic characteristics of the intact epithelium, including apical microvilli, apical tight junctions, and a moderately interdigitated lateral intercellular space. The cells also retained the functional properties of the epithelium. The monolayer responded to addition of isoproterenol with the characteristic changes in cellular electrical properties expected for stimulation of C1 secretion: isoproterenol increased transepithelial voltage, depolarized apical membrane voltage, and decreased both transepithelial resistance and the ratio of apical-to-basolateral membrane resistance. Examination of the cellular response to ion substitutions and inhibitors of C1 secretion indicate that the cultured monolayers retain the same cellular mechanisms of ion transport as the intact epithelium. Thus, primary cultures of tracheal epithelium may provide a useful preparation for future studies of the mechanism and regulation of C1 secretion by airway epithelia.

Electro-osmosis and the reabsorption of fluid in renal proximal tubules.

The lateral intercellular spaces (LIS) are believed to be the final common pathway for fluid reabsorption from the renal proximal tubule. We postulate that electrogenic sodium pumps in the lateral membranes produce an electrical potential within the LIS, that the lateral membranes bear a net negative charge, and that fluid moves parallel to these membranes because of Helmholtz-type electro-osmosis, the field-induced movement of fluid adjacent to a charged surface. Our theoretical analysis indicates that the sodium pumps produce a longitudinal electric field of the order of 1 V/cm in the LIS. Our experimental measurements demonstrate that the electrophoretic mobility of rat renal basolateral membrane vesicles is 1 micron/s per V/cm, which is also the electro-osmotic fluid velocity in the LIS produced by a unit electric field. Thus, the fluid velocity in the LIS due to electro-osmosis should be of the order of 1 micron/s, which is sufficient to account for the observed reabsorption of fluid from renal proximal tubules. Several experimentally testable predictions emerge from our model. First, the pressure in the LIS need not increase when fluid is transported. Thus, the LIS of mammalian proximal tubules need not swell during fluid transport, a prediction consistent with the observations of Burg and Grantham (1971, Membranes and Ion Transport, pp. 49-77). Second, the reabsorption of fluid is predicted to cease when the lumen is clamped to a negative voltage. Our analysis predicts that a voltage of -15 mV will cause fluid to be secreted into the Necturus proximal tubule, a prediction consistent with the observations of Spring and Paganelli (1972, J. Gen. Physiol., 60:181).

1610 MODEL OF ACUTE CYCLOSPORINE-INDUCED NEPHROTOXI CITY (ACIN) IN THE GROWING RAT

Immunosuppression with cyclosporine (CYC) results in an early, acute, reversible reduction in renal hemodynamlcs. In an attempt to establish a model for investigating the functional and morphologic effects of ACIN at the single nephron level, renal micropuncture (MP) and ultra-structural (EM) studies were performed in growing male rats (100-140 g). Pair-caged animals were treated with IM injections of CYC (10 or 25 mg/kg/d) or olive oil for 1 week. Daily weight gain was reduced in both treated groups (-32%, n=24, and -44%, n=6, p<.01). Inactin anesthesia (110-120 mg/kg IP) and the hydropenic (HYP) state were maintained throughout MP. Rats given 25 mg/kg CYC were hypothermic and hypotensive; few survived the surgical preparation for MP. Kidneys appeared grossly abnormal with no superficial tubular flow. Kidney gloraerular filtration rate (GFR) was less in the 10 mg/kg CYC treated rats (0.39 ± .07 S.E. ml/min/100 g, n=4), than in a separate group of HYP controls of the same weight (0.56 ± .04 ml/min/100 g, n=5, p<.005). Proximal single nephron GFR was also lower (24.2 ± 1.8 nl/min, n=4) than the controls (34.0 ± 2.6 nl/min, n=9, p<.001). EM studies performed in a group of identically treated rats showed distal tubules with widely dilated lateral intercellular spaces. Proximal tubules had increased intracellular vacuoles. These results agree with observations in human ACIN and demonstrate that this is a useful animal model of ACIN.

Passive water flows driven across the isolated rabbit ileum by osmotic, hydrostatic and electrical gradients.

Water flows generated by osmotic and hydrostatic pressure and electrical currents were measured in sheets of isolated rabbit ileum at 20 degrees C. Flows across the mucosal and serosal surfaces were monitored continuously by simultaneous measurement of tissue volume change (with an optical lever) and net water flows across one surface of the tissue (with a capacitance transducer). Osmotic gradients were imposed across the mucosal and serosal surfaces of the tissue separately, using probe molecules of various sizes from ethanediol (68 Da) to dextrans (161 000 Da). Flows across each surface were elicited with very short delay. The magnitudes of the flows were proportional to the osmotic gradient and related to the size of the probe molecule. Osmotic flow across the mucosal surface was associated with streaming potentials which were due to electro-osmotic water flow. The mucosal surface is a heteroporous barrier with narrow (0.7 nm radius, Lp (hydraulic conductivity) = (7.6 +/- 1.6) X 10(-9) cm s-1 cmH2O-1) cation-selective channels in parallel with wide neutral pores (ca. 6.5 nm radius, Lp = (2.3 +/- 0.2) X 10(-7) cm s-1 cmH2O-1) which admit large pressure-driven backflows from the submucosa to the lumen. There is additional evidence for a further set of narrow electroneutral pores less than 0.4 nm radius with Lp less than 7 X 10(-9) cm s-1 cmH2O-1. The serosal surface has neutral pores of uniform radius (ca. 6.5 nm), Lp = (7.6 +/- 1.6) X 10(-8) cm s-1 cmH2O-1. Hypertonic serosal solutions (100 mM-sucrose) cause osmotic transfer of fluid from isotonic mucosal solutions into the submucosa, expand it, and elevate the tissue pressure to 19.6 +/- 3.2 cmH2O (n = 4). Conversely, hypertonic mucosal solutions (100 mM-sucrose) draw fluid out of the submucosa in the presence of isotonic serosal solutions, collapse the submucosa, and lower the tissue pressure to -87.7 +/- 4.6 cmH2O (n = 5). Water flows coupled to cation movement could be generated across the mucosal surface in both directions by brief direct current pulses. The short latency of onset and cessation of flow (less than 2 s), absence of polarization potentials, and high electro-osmotic coefficients (range 50-520 mol water F-1), together with the presence of streaming potentials during osmotically generated water flows indicate electro-osmotic water flow through hydrated channels in the tight junctions and/or lateral intercellular spaces.(ABSTRACT TRUNCATED AT 400 WORDS)

Protein and lipid absorption in the intestinal mucosa of adult lampreys (Petromyzon marinus L.) following induced feeding

Young adult lampreys (Petromyzon marinus) were force-fed either on polyunsaturated lipid or on a protein tracer following a period of starvation to follow the absorption and intracellular transport of these substances in the intestine. Type A and B absorptive cells of the diverticulum, anterior intestine, and transition zone are the principal cells of lipid absorption and transport, whereas protein absorption is confined to caveolated cells of the posterior intestine and hindgut. Lipoprotein, corresponding to very low density lipoprotein (VLDL), becomes localized in smooth and rough endoplasmic reticulum, smooth tubules, and in the Golgi saccules and vesicles, while lipid droplets are localized in the cytoplasmic matrix. The localization of VLDL in the lateral intercellular space and extracellular space, including the submucosal connective tissue and the lumina of the capillaries, are believed to be an expression of the manner in which lipid is absorbed and transported in the anterior segments of the lamprey intestine. Protein absorption in caveolated cells follows the pattern noted in cells of similar function in other fishes and involves pinocytotic vesicles, a tubulovesicular network, heterophagic vacuoles, and likely a lysosomal system for degradation and condensation of the endocytosed material. The presence of regions specialized for lipid and protein absorption in the intestine of adult lampreys is comparable to the situation in many other fishes.

Electrophysiological studies on lateral intercellular spaces ofNecturus gallbladder epithelium

The lateral intercellular spaces of Necturus gallbladder epithelium were punctured with double-barrelled ion selective microelectrodes in order to determine the ion concentrations of lateral space fluid and the contribution of the lateral spaces to transepithelial resistance. Neither under control conditions, nor after diluting the bathing fluids to increase the rate of volume absorption, nor during passage of direct current of 200 microA/cm2, were any reliable concentration differences observed between lateral space fluid and external bathing fluids. These observations suggest that water can follow salt transport without requiring osmotic concentration gradients of greater than 1 or 2 mosmol/l and indicate that recently observed high values of water permeability must still be considered as underestimates. After developing a test to recognize and exclude leaky punctures, the contribution of the lateral spaces to transepithelial resistance could be determined. It amounted to around 29%. This value agrees well with results from recent impedance measurements which were performed under control conditions in the same preparation.

The Lateral Intercellular Spaces in the Endolymphatic Sac. A Pathway for Fluid Transport?

The lateral intercellular spaces (LIS) in the epithelium of the endolymphatic sac in different mammals were studied using light and transmission electron microscopy. The reason for the study was that widened LIS are known to occur in fluid transporting epithelia, where they may reflect transepithelial flux of water and solutes. Increased knowledge about LIS may lead to further insight into the mechanisms of endolymph resorption in the endolymphatic sac. The effects of various fixatives, fixation methods and osmolality, and also of surgical labyrinthectomy, on the ultrastructure of LIS were investigated. Widened LIS regularly occurred in the mammalian endolymphatic sac and seem to reflect an in vivo condition. It is thought that LIS may form a pathway for transepithelial water flow in the endolymphatic sac. A hypothetical model of the function of LIS during transepithelial fluid movement in the sac is presented.

Transport by epithelia with compliant lateral intercellular spaces: asymmetric oncotic effects across the rat proximal tubule.

Mathematical models of the proximal tubule are considered in which the lateral intercellular spaces distend in response to increased interstitial pressures and basal outlet permeabilities increase as a result of interspace widening. An approximate analytical model of the interspace reveals the possibility that such compliance may introduce an asymmetry to the effect of protein oncotic forces on transepithelial volume flow. Peritubular oncotic forces close the interspace, enhance interspace hypertonicity, and thus substantially increase volume reabsorption (enhanced intraepithelial solute-solvent coupling). The model also predicts a decline in epithelial water permeability (Lp), salt reflection coefficient, and salt permeability, with the application of peritubular protein. When parameters are chosen so as to represent the rat proximal tubule, the predicted effect on solute permeability is comparable to the observed changes in electrical resistance of the epithelium. However, when the luminal solution is slightly hypotonic to blood and proximal reabsorption has become isosmotic, the models show relatively small protein effects, which are dependent upon cell and tight junction permeabilities and are little influenced by interspace compliance. The capability of such models to represent the peritubular protein enhancement of isosmotic salt and water reabsorption by the proximal tubule in vivo is questioned.

Endocytosis in epithelial cells lining the rete testis of the rat.

The endocytic activity of the low cuboidal cells lining the rete testis was analyzed by electron microscopy following injection of various tracers into the lumen of these anastomotic channels. At 1 and 5 minutes after injection, cationic ferritin (CF) and concanavalin A-ferritin (Con A) were seen bound to the apical plasma membrane and to the membrane of subjacent vesicles or invaginations connected to this apical membrane. At 30 and 60 minutes, these tracers were found in intracytoplasmic vesicles and in vesicles connected to the lateral or basal plasma membrane as well as in the lateral intercellular space and in the lamina lucida of basal lamina. At 30 minutes, CF and Con A also appeared in the matrix of pale multivesicular bodies while at 1 hour dense multivesicular bodies were labeled. At 2 hours and later time intervals, the tracers accumulated in dense granules identified as lysosomes. Native ferritin (NF), concanavalin A-ferritin in presence of alpha-methyl-D-mannoside, and horseradish peroxidase or albumin bound to colloidal gold were all to be incorporated by the lysosomal system of these epithelial cells, as just described for CF and Con A, but these various tracers were not bound to the apical plasma membrane or to the membrane of cytoplasmic vesicles, nor were they found in the intercellular spaces or the lamina lucida at the base of the cells. Thus, the epithelial cells of the rete testis do not appear to be only involved in the uptake of substances from the lumen and their disposal by the lysosomal system, but also appear to contribute to the transport of certain macromolecules from the lumen to the laterobasal surfaces of the cells. These cells may thus play a role in determining the composition of the rete testis fluid.

Differentiation of the segmented tubular nephron and excretory duct during lamprey metamorphosis

The tubular portion of the adult lamprey nephron differentiates into various morphologically distinct segments during the seven stages of metamorphosis. At stage 1, a rudimentary nephron unit (RNU) originates from a nephrogenic mass attached to the peritoneum and elongates to become associated at its distal end with the archinephric (excretory) duct. The rapidly dividing cells show little sign of differentiation. Separation of the RNU from the peritoneum in stage 2 is first accompanied by widening of lateral intercellular spaces and then eventually by the formation of a small lumen at the proximal end. Cells surrounding a portion of this lumen show early signs of ciliogenesis and hence are located in the position of the presumptive neck segment. The distal end of the stage 2 RNU contains an electron-dense material in an enlarged intercellular space and the cells make non-junctional contact with the archinephric duct. The larval cells of the latter structure are either lost during degeneration and desquamation or are transformed into adult cells. By the end of stage 3 replication of basal bodies and formation of cilia has resulted in the appearance of a ciliated neck segment while numerous microvilli extending into a narrow lumen mark the position of the proximal segment. Furthermore, most of the distal portion of the newly formed tubule possesses a lumen containing a compact mass of material likely secreted by the cells during lumen formation. This presumptive distal segment terminates in the transforming epithelium of the archinephric duct as a presumptive connecting or collecting segment. Fully differentiated neck and proximal segments are present in stage 4 with the latter having a complete apparatus for endocytosis but the distal segment shows only early signs of differentiation of the smooth tubular network. This network appears to arise as an expanded surface area of the plasma membrane. Stage 5 is characterized by elongation of the proximal portion of the tubule, expansion of the tubular network in distal cells, and the identification of a collecting segment. The last two stages show no further major differentiation of the segments. The early differentiation of tubular segments in the adult lamprey nephron likely reflects the immediate physiological needs of this organism as the larval kidney undergoes regression. The potential of the developing kidney of adult lampreys as a source of information on morphogenesis of microvilli, the endocytotic apparatus, mitochondria, and a smooth tubular network is discussed. It appears that the archinephric duct and non-nephrogenic mesenchyme may play some role in tubulogenesis in lampreys.