The Ukrainian brook lamprey Eudontomyzon mariae is the most widespread lamprey species in eastern Europe. Although E. mariae is generally considered a derivative of Eudontomyzon danfordi, an exclusively freshwater parasitic species, it has alternatively been suggested that it was recently derived from a now extinct anadromous Black Sea ancestor. Several non-parasitic lampreys and the landlocked sea lamprey, which have recently evolved from anadromous ancestors, still develop a seawater-type mitochondria-rich cell (SW-MRC) in their gills. In contrast, this cell type is not present in the gills of either Lampetra aepyptera, a non-parasitic lamprey of ancient origin, or the parasitic Ichthyomyzon unicuspis and I. castaneus that likewise have long evolutionary histories in fresh water. Eudontomyzon mariae from the Vistula River in the Baltic River basin does not possess SW-MRC, which is inconsistent with a recent origin from an anadromous ancestor. Mitochondrial DNA sequence data were thus used to infer the relationship between different populations of E. mariae and E. danfordi, and to reconstruct the transition from anadromy to freshwater residency. The results suggest that E. mariae evolved independently in the Baltic, Black, and Caspian Sea basins, and not recently from an anadromous ancestor. Although E. mariae in the Danube River may have arisen relatively recently from E. danfordi (differing by 0.7–1.1% in cytochrome b gene sequence), other E. mariae populations (including in the Vistula River) are genetically closer (0.6%) to the hypothetical ancestor of both E. mariae and E. danfordi. That ancestor was probably a freshwater resident, since SW-MRCs are not rapidly lost following confinement in fresh water.

Although confined to fresh water, non‐parasitic species of lampreys and the landlocked parasitic sea lamprey, all of which were derived relatively recently from anadromous ancestors, still develop chloride cells, whose function in their ancestors was for osmoregulation in marine waters during the adult parasitic phase. In contrast, such cells are not developed by the non‐parasitic least brook lamprey Lampetra aepyptera, which has been separated from its ancestor for >2 million years, nor by the freshwater parasitic species of the genus Ichthyomyzon. The length of time that a non‐parasitic species or landlocked parasitic form or species has spent in fresh water is thus considered the overriding factor determining whether chloride cells are developed by those lampreys.

TonEBP/NFAT5 is a major regulator of the urinary concentrating process and is essential for the osmoadaptation of renal medullary cells. Focal adhesion kinase (FAK) is a mechanosensitive non-receptor protein tyrosine kinase expressed abundantly in the renal medulla. Since osmotic stress causes cell shrinkage, the present study investigated the contribution of FAK on TonEBP/NFAT5 activation. Osmotic stress induced time-dependent activation of FAK as evidenced by phosphorylation at Tyr-397, and furosemide reduces FAK Tyr-397 phosphorylation in the rat renal medulla. Both pharmacological inhibition of FAK and siRNA-mediated knockdown of FAK drastically reduced TonEBP/NFAT5 transcriptional activity and target gene expression in HEK293 cells. This effect was not mediated by impaired nuclear translocation or by reduced transactivating activity of TonEBP/NFAT5. However, TonEBP/NFAT5 abundance under hypertonic conditions was diminished by 50% by FAK inhibition or siRNA knockdown of FAK. FAK inhibition only marginally reduced transcription of the TonEBP/NFAT5 gene. Rather, TonEBP/NFAT5 mRNA stability was diminished significantly by FAK inhibition, which correlated with reduced reporter activity of the TonEBP/NFAT5 mRNA 3' untranslated region (3'-UTR). In conclusion, FAK is a major regulator of TonEBP/NFAT5 activity by increasing its abundance via stabilization of the mRNA. This in turn, depends on the presence of the TonEBP/NFAT5 3'-UTR.

Background: The exoenzyme carbonic anhydrase IX is strongly expressed in tumours. In normal tissues, CA IX is expressed during embryogenesis but downregulated postnatally, except in the stomach. Aim and Methods: We studied the cellular differentiation pattern and acid secretory capacity in gastric mucosa from CAIX KO and WT mice at age 8 days to 1,3,6,9,12 months. Surface pH and mucus layer thickness, as well as the ability of the surface epithelial cells to withstand a luminal acid load was assessed in young adult CAIX KO and WT mice by in vivo two photon microscopy. Results: CAIX KO gastric mucosa developed a unique phenotype with a gradual expansion of the surface cell zone, and regression of the parietal cell zone to the lower gland area. Concomitantly, the acid secretory capacity, which was normal at young age, declined. HCO3‐ output into the lumen and mucus layer thickness were not significantly different between CAIX KO and WT. However, the ability of the surface cells to withstand a luminal acid load, as well as to maintain a more alkaline surface pH than the luminal pH, was significantly impaired in CAIX KO compared to WT stomach. Conclusion: We speculate that the CAIX at the basolateral membrane of the gastric surface cells rapidly converts extruded protons with blood‐borne HCO3‐ to CO2 and thus augments interstitial buffer capacity, surface cell pHi regulation, and maintenance of the pH microclimate in the mucus layer.Grant Funding Source: Supported by DFG Se 13‐4 and 9‐6 to Ursula Seidler

This paper provides the first description of the cellular composition of the gill epithelium of feeding adults of Ichthyomyzon unicuspis Hubbs and Trautman, 1937 (silver lamprey), a parasitic species of lamprey that is confined to fresh water. The surface layer of this epithelium consists solely of pavement cells and intercalated mitochondria-rich cells, which are the only cell types found in all freshwater stages of lampreys and thus considered responsible for the uptake of Na+ and Cl– in hypotonic environments. This epithelium does not contain, however, the chloride cells present during the marine parasitic phase of anadromous lamprey species, such as Petromyzon marinus L., 1758 (sea lamprey), and which are responsible for secreting excess Na+ and Cl–. The absence of this cell type in parasitic adults of I. unicuspis also differs from its presence in parasitic adults of landlocked P. marinus and metamorphosing individuals of the exclusively freshwater nonparasitic species Lethenteron appendix (DeKay, 1842) (American brook lamprey), and which thus reflects the retention of a cell type that was crucial for osmoregulation during the marine phase of their respective anadromous parasitic ancestors. The absence of chloride cells in I. unicuspis is consistent with the hypothesis that Ichthyomyzon, which is at or close to the base of the phylogenetic tree for Northern Hemisphere lampreys (Petromyzontidae), evolved in fresh water or has been confined to fresh water for a very long period.

Several lamprey species form pairs, comprising an anadromous parasitic species and a derivative nonparasitic species that neither leaves fresh water nor feeds as an adult. This paper provides the first description of the radical changes undergone by the cellular composition of the gill epithelium during the major phases in the life cycle of a nonparasitic lamprey (American brook lamprey, Lethenteron appendix (DeKay, 1842) (= Lampetra appendix (DeKay, 1842)) and discusses their potential functional and evolutionary significance. The gill epithelium of the larva of L. appendix contains ammocoete mitochondrion-rich cells (AMRCs), intercalated mitochondrion-rich cells, and pavement cells, as does that of the larva of anadromous parasitic species which likewise lives in fresh water. By the completion of metamorphosis, the AMRCs have disappeared and well-developed chloride cells have been produced, the latter cell type being essential for osmoregulation by its closely related anadromous species in hypertonic environments. By the attainment of sexual maturity, the chloride cells have been lost. Such changes in the timing of chloride cell representation could help account for the ability of some metamorphosing, but not mature individuals of another nonparasitic species (Lampetra planeri (Bloch, 1784)), to osmoregulate in up to 70% of seawater. The well-developed chloride cells in the nonparasitic L. appendix represent the retention of an ancestral character.

During antidiuresis, cell survival in the renal medulla requires cyclooxygenase-2 (COX-2) activity. We have recently found that prostaglandin E2 (PGE(2)) promotes cell survival by phosphorylation and, hence, inactivation of the pro-apoptotic protein Bad during hypertonic stress in Madin-Darby canine kidney (MDCK) cells in vitro. Here we determine the role of COX-2-derived PGE(2) on phosphorylation of Bad and medullary apoptosis in vivo using COX-2-deficient mice. Both wild-type and COX-2-knockout mice constitutively expressed Bad in tubular epithelial cells of the renal medulla. Dehydration caused a robust increase in papillary COX-2 expression, PGE(2) excretion, and Bad phosphorylation in wild-type, but not in the knockout mice. The abundance of cleaved caspase-3, a marker of apoptosis, was significantly higher in papillary homogenates, especially in tubular epithelial cells of the knockout mice. Knockdown of Bad in MDCK cells decreased tonicity-induced caspase-3 activation. Furthermore, the addition of PGE(2) to cells with knockdown of Bad had no effect on caspase-3 activation; however, PGE(2) caused phosphorylation of Bad and substantially improved cell survival in mock-transfected cells. Thus, tonicity-induced COX-2 expression and PGE(2) synthesis in the renal medulla entails phosphorylation and inactivation of the pro-apoptotic protein Bad, thereby counteracting apoptosis in renal medullary epithelial cells. Kidney International (2011) 80, 938-945; doi:10.1038/ki.2011.199; published online 29 June 2011

Dendritic cells (DCs) are central for the induction of T cell immunity and tolerance. Fundamental for DCs to control the immune system is their differentiation from precursors into various DC subsets with distinct functions and locations in lymphoid organs and tissues. In contrast to the differentiation of epidermal Langerhans cells (LCs) and their seeding into the epidermis, LC maturation, turnover, and MHC class II Ag presentation capacities are strictly dependent on the presence of Dicer, which generates mature microRNAs (miRNAs). Absence of miRNAs caused a strongly disturbed steady-state homeostasis of LCs by increasing their turnover and apoptosis rate, leading to progressive ablation of LCs with age. The failure to maintain LCs populating the epidermis was accompanied by a proapoptotic gene expression signature. Dicer-deficient LCs showed largely increased cell sizes and reduced expression levels of the C-type lectin receptor Langerin, resulting in the lack of Birbeck granules. In addition, LCs failed to properly upregulate MHC class II, CD40, and CD86 surface molecules upon stimulation, which are critical hallmarks of functional DC maturation. This resulted in inefficient induction of CD4 T cell proliferation, whereas Dicer-deficient LCs could properly stimulate CD8 T cells. Taken together, Dicer-dependent generation of miRNAs affects homeostasis and function of epidermal LCs.

Three kinds of epithelial cells comprise the surfaces of the gill filaments and lamellae of larval lampreys (ammocoetes): ammocoete mitochondria-rich cells (AMRCs), intercalated mitochondria-rich cells (IMRCs) and pavement cells. Selected characteristics of these cell types in ammocoetes of Geotria australis held in distilled water and in 10% sea water were compared using an ultrastructural stereological approach to determine which of those cell type(s) respond to exposure to an ion-deficient environment in a manner that indicates that they are involved in ion uptake. Particular focus was placed on the enigmatic AMRC, which comprises ca 60% of the cells and contains numerous mitochondria. The mean percentage contributions of both AMRCs and pavement cells to the total number of the three cell types in the two experimental groups were not significantly different, whereas that of IMRCs was >7% in distilled water and <1% in 10% sea water (P < 0.001). Furthermore, the mean apical surface areas of neither AMRCs nor pavement cells differed significantly between the two experimental groups, whereas that of IMRCs was nearly 3-fold greater in distilled water than in 10% sea water. The volume densities and size of mitochondria in AMRCs did not differ between the two exposure regimes. The above comparisons provide no indications that the uptake of Na(+) and Cl(-) in the gill epithelium of ammocoetes involves either the AMRC or pavement cell but, when considered in conjunction with data on ion-transporting cells in other vertebrates, they are consistent with the conclusion that the IMRC plays a crucial role in this process.

Cyooxygenase-2 (COX-2)-derived PGE2 is critical for the integrity and function of renal medullary cells during antidiuresis. The present study extended our previous finding that tonicity-induced COX-2 expression is further stimulated by the major COX-2 product PGE2 and investigated the underlying signaling pathways and the functional relevance of this phenomenon. Hyperosmolality stimulated COX-2 expression and activity in Madin-Darby canine kidney (MDCK) cells, a response that was further increased by PGE2-cAMP signaling, suggesting the existence of a positive feedback loop. This effect was diminished by AH-6809, an EP2 antagonist, and by the PKA inhibitor H-89, but not by AH-23848, an EP4 antagonist. The effect of PGE2 was mimicked by forskolin and dibutyryl-cAMP, suggesting that the stimulatory effect of PGE2 on COX-2 is mediated by a cAMP-PKA-dependent mechanism. Accordingly, cAMP-responsive element (CRE)-driven reporter activity paralleled the effects of PGE2, AH-6809, AH-23848, H-89, forskolin, and dibutyryl-cAMP on COX-2 expression. In addition, the stimulatory effect of PGE2 on tonicity-induced COX-2 expression was blunted in cells transfected with dominant-negative CRE binding (CREB) protein, as was the case in a COX-2 promoter reporter construct in which a putative CRE was deleted. Furthermore, PGE2 resulted in PKA-dependent phosphorylation of the pro-apoptotic protein Bad at Ser155, a mechanism that is known to inactivate Bad, which coincided with reduced caspase-3 activity during osmotic stress. Conversely, pharmacological interruption of the PGE2-EP2-cAMP-PKA pathway abolished Ser155 phosphorylation of Bad and blunted the protective effect of PGE2 on cell survival during osmotic stress. These observations indicate the existence of a positive feedback loop of PGE2 on COX-2 expression during osmotic stress, an effect that apparently is mediated by EP2-cAMP-PKA signaling, and that contributes to cell survival under hypertonic conditions.

In the renal medulla, cyclooxygenase (COX)-2 is induced by osmotic stress as present in this kidney region during antidiuresis. Increasing evidence suggests that EGF receptor ( EGFR) signaling is involved in this process. The aim of the present study was to examine the mechanisms responsible for COX-2 expression and PGE(2) production during hypertonic conditions and to identify potential autocrine/paracrine EGFR ligands. Immunohistochemisty and Western blot analysis revealed abundant expression of the pro-EGFR ligand pro-transforming growth factor (TGF)-alpha in renal medullary cells in vivo and in cultured Madin-Darby canine kidney cells. In Madin-Darby canine kidney cells, hypertonicity rapidly increased TNF-alpha converting enzyme (TACE)-dependent ectodomain shedding of pro-TGF-alpha; phosphorylation of EGFR, p38, and ERK1/2; expression of COX-2; and production of PGE2. Conversely, TACE inhibition prevented TGF-alpha release; EGFR, p38, and ERK1/2 activation; and COX-2 expression. Furthermore, cell survival was reduced substantially, a response that could be reversed by the addition of PGE2. Simultaneous addition of recombinant TGF-alpha during TACE inhibition restored EGFR and MAPK phosphorylation, COX-2 expression, PGE2 production, and cell survival during osmotic stress. These results indicate that hypertonicity induces TACE-mediated ectodomain shedding of pro-TGF-alpha, which subsequently activates COX-2 expression in an autocrine/paracrine fashion, via EGFR and MAPKs. We conclude that tonicity-induced TGF-alpha release is required for COX-2 expression, PGE2 synthesis, and survival of renal medullary cells during osmotic stress.

Focal adhesion kinase (FAK), a non-receptor tyrosine kinase, plays important roles in cell migration, cell proliferation and cell survival. Because these processes participate in the restoration of tubular integrity in renal ischaemia and reperfusion, FAK expression and phosphorylation at Tyr-397, the latter indicative of its activity, were examined in the different kidney zones by Western blot analysis and immunohistochemistry. Expression and phosphorylation of FAK were also studied in Madin-Darby canine kidney (MDCK) and medullary thick ascending limb (mTAL) cells after ATP depletion and repletion. In control rat kidneys, FAK expression in outer and inner medulla exceeded that in cortex, and phosphorylation of FAK at Tyr-397 was most pronounced in the inner medulla. Although this expression pattern was not affected by 20 (40, 60)-min ischaemia and 20 (40, 60)-min ischaemia followed by 60-min or 24-h reperfusion, FAK phosphorylation was significantly reduced in all kidney zones immediately after ischaemia, but increased during reperfusion, exceeding control values in the outer and inner medulla. ATP depletion and repletion of MDCK and mTAL cells were associated with a decrease in FAK phosphorylation during ATP depletion, followed by an increase during repletion. Rephosphorylation of FAK after ATP repletion was enhanced by N -acetylcysteine, a reactive oxygen species scavenger. ATP depletion disrupted focal adhesions in MDCK cells. Their reformation after ATP repletion paralleled the increase in FAK phosphorylation. These findings suggest an essential role for FAK-signalling during renal ischaemia and early reperfusion.

SUMMARY Lampreys, one of the only two surviving groups of agnathan (jawless)vertebrates, contain several anadromous species that, during their life cycle,thus migrate from fresh to seawater and back to freshwater. Lampreys have independently evolved the same overall osmoregulatory mechanisms as the gnathostomatous (jawed) and distantly related teleost fishes. Lamprey gills thus likewise play a central role in taking up and secreting monovalent ions. However, the ultrastructural characteristics and distribution of their epithelial cell types [ammocoete mitochondria-rich (MR) cell, intercalated MR cell, chloride cell and pavement cell] differ in several respects from those of teleosts. The ultrastructural characteristics of these cells are distinctive and closely resemble those of certain ion-transporting epithelia in other vertebrates, for which the function has been determined. The data on each cell type, together with the stage in the life cycle at which it is found, i.e. whether in fresh or seawater, enable the following proposals to be made regarding the ways in which lampreys use their gill epithelial cells for osmoregulating in hypo- and hypertonic environments. In freshwater, the intercalated MR cell takes up Cl– and secretes H+,thereby facilitating the uptake of Na+ through pavement cells. In seawater, the chloride cell uses a secondarily active transcellular transport of Cl– to provide the driving force for the passive movement of Na+ through leaky paracellular pathways between these cells.

Yersinia enterocolitica cross the intestinal epithelium via translocation through M cells, which are located in the follicle-associated epithelium (FAE) of Peyer's patches (PP). To investigate the molecular basis of this process, studies were performed using a recently developed in vitro model, in which the enterocyte-like cell line Caco-2 and PP lymphocytes are co-cultured in order to establish FAE-like structures including M cells. Here, we demonstrate that Y. enterocolitica does not adhere significantly to the apical membrane of differentiated enterocyte-like Caco-2 cells that express binding sites for Ulex europaeus agglutinin (UEA)-1. In contrast, Y. enterocolitica adhered to, and was internalized by, cells that lacked UEA-1 binding sites and displayed a disorganized brush border. These cells were considered to be converted to M-like cells. Further analysis revealed that part of these cells expressed beta 1 integrins at their apical surface and, as revealed by comparison of wild-type and mutant strains, interacted with invasin of Y. enterocolitica. Consistently, anti-beta 1 integrin antibodies significantly inhibited internalization of inv-expressing yersiniae. Experiments with Yersinia mutant strains deficient in YadA or Yop secretion revealed that these virulence factors play a minor role in this process. After internalization, yersiniae were transported within LAMP-1-negative vacuoles to, and released at, the basal surface. Internalization and transport of yersiniae was inhibited by cytochalasin D, suggesting that F-actin assembly is required for this process. These results provide direct evidence that expression of beta 1 integrins at the apical surface of M cells enables interaction with the invasin of Y. enterocolitica, and thereby initiates internalization and translocation of bacteria.

The distribution and ultrastructure of the mitochondria-rich (MR) cells in the gills of larval (ammocoetes) and adult lampreys (Petromyzon marinus and Geotria australis) have been studied. One type of MR cell, which is found only in ammocoetes, occurs in groups on and between gill lamellae. Freeze-fracture replicas show that the apical membrane of this ammocoete MR cell contains globular particles. The second type of MR cell, which is present in both ammocoetes and adults in freshwater, is located between lamellae and at the base of the filament. This cell usually occurs singly and is typically intercalated between ammocoete MR cells in larval lampreys and between pavement cells and pavement and chloride cells in adult lampreys. It contains rod-shaped particles in either the apical membrane (subtype A) or, far less frequently, the lateral membrane (subtype B) and in membranes of cytoplasmic vesicles and tubules. These features characterize this intercalated MR cell as a member of a group of MR cells that are also found in urinary epithelia of tetrapods and the amphibian epidermis, where they are involved in H+ and HCO3- secretion. Because this type of MR cell disappears when the young adult lamprey enters the sea and reappears immediately after the fully grown adult re-enters freshwater on its spawning run, it is presumably essential for osmoregulation in freshwater. On the basis of electrophysiological studies on frog skin, it is proposed that the subtype A of the branchial intercalated MR cell of lampreys provides the driving force for the Na+ uptake by active H+ secretion. By analogy with urinary epithelia, the subtype B cells may exchange Cl- for HCO3-.

Thin-section and freeze-fracture electron microscopy have been used to determine the types of junctions that are present in the notochord of adult lampreys and thereby propose ways in which the functional properties of the notochord are integrated. The notochord consists of a fibrous sheath, which surrounds a single layer of small 'epithelial' cells and a central core of large vacuolated cells. The epithelial and vacuolated notochordal cells are all linked by numerous desmosomes, which are the predominant type of intercellular junction in the notochord. Furthermore, the notochordal epithelium is linked to the inner surface of the notochordal sheath by many hemidesmosomes, which are associated with patches of basal lamina-like material. The above linkages, produced by desmosomes and hemidesmosomes, together with the tensile strength of the notochordal sheath, must he crucial for maintaining the structural integrity of the notochord during the sinuous swimming movements exhibited by lampreys. The communicating (gap) junctions, that are present between vacuolated cells, would facilitate metabolic coupling between these cells. Zonulae occludentes ('tight junctions') were not found between notochordal epithelial cells, indicating that there is no effective paracellular barrier to the diffusion of macromolecules, ions and small polar solutes at the outermost layer of the notochordal cells.

The gills of hagfishes form lens-shaped pouches, rather than holobranches that characterize those of lampreys and gnathostome fishes. Their internal surface is enlarged by radial folds, which extend between the medial and lateral walls of the pouch and contain the branchial microcirculation. Blood and water flow through the gills are arranged in a countercurrent system, thereby facilitating gas exchange.The circulation of the hagfish gills consists of an arterio-arterial and an arteriovenous component. The arterio-arterial circulation connects the afferent and efferent branchial arteries. The arteriovenous circulation comprises an intrabranchial sinusoid system, which is supplied by the arterio-arterial circulation through arteriovenous anastomoses and connected to the peribranchial sinus through gaps in the muscle layer of the gill pouch.The majority of the epithelial surface is formed by mucus-secreting pavement cells. In the lateral half of the gill pouch, mitochondria-rich (MR) cells are intercalated between the pavement cells, which are characterized by an extensive amplification of the basolateral cell membrane and, in freeze-fracture replicas, by assemblies of linear arrays of particles and fibrils in the apical cell membrane. Since the MR cells show strong histochemical reactions for both Na+/K+-ATPase and carbonic anhydrase, they may be engaged in ion transport and/or acid-base regulation.

The M-cells at four locations (Peyer's patches, sacculus rotundus, cecal lymphoid patch, and appendix) of gut-associated lymphoid tissue (GALT) in the rabbit ileum and cecum were studied by light microscopy, lectin histochemistry, immunohistochemistry, and thin-section, freeze fracture and scanning electron microscopy. M-cells differ from adjacent enterocytes in a less regularly shaped apical surface, a deep invagination of the basolateral cell surface that engulfs intraepithelial lymphoid cells, the ability to transcytose antigens, and the presence of vimentin intermediate filaments in addition to cytokeratins. Differences between M-cells of Peyer's patches and the cecal patch include the presence of a distinct lectin-labelling pattern and a deep invagination of the apical surface that bypasses a pronounced terminal web in cecal M-cells. M-cells in the dome epithelia of sacculus rotundus and appendix, but not in Peyer's and cecal patches are linked to adjacent enterocytes by extended fasciae occludentes on the lateral membrane in addition to the belt-like zonulae occludentes regularly present in simple epithelia. The presence of these fasciae occludentes correlates with the appearance of regularly shaped clusters of lymphoid cells enclosed by the pocket-like invagination of the basal M-cell surface at these locations. The differences between M-cells in Peyer's and cecal patches might be related to differences in the gut content between the small and Large intestines such as the microbial population, whereas the characteristics of M-cells in the sacculus rotundus, which is located in the ileum and the appendix as a part of the cecum, are obviously not related to this situation.

Scanning electron microscopy (SEM) was used to study the changes that occur in the morphological relationships between chloride and pavement cells in the gills during acclimation of young adult lampreys to seawater. Because chloride cells are located predominantly between lamellae and are thus obscured from view, the lamellae were removed with the use of a micromanipulator installed in a SEM. In gills of animals maintained in river water, chloride cells could then be seen to be dislike and typically to form single rows between successive lamellae. After acclimation to seawater, the apical surfaces of chloride cells lose their microvilli and change in shape from small circles to rectangles that extend the full width between successive lamellae. These changes result in an increase in the length of the paracellular pathway between chloride cells. Previous work has shown that the number of strands of the zonulae occludentes sealing this pathway declines under these conditions. This presumably leads to an increase in paracellular permeability of the gill epithelium, thereby providing the low-resistance paracellular shunt required for the passive movement of sodium into the environment during osmoregulation in seawater. The above changes are reversed by transfer of lampreys downward to 10% seawater.

Larvae of the two southern hemisphere lamprey genera, Mordacia and Geotria, possess one and two intestinal diverticula, respectively, each originating at the oesophageal-intestinal junction. These diverticula comprise an inner layer of simple columnar epithelium composed solely of zymogen and mucous cells, a middle layer consisting mainly of a blood sinus, and an outer serosa layer covered by a simple squamous epithelium (mesothelium). The inner surface is highly folded only in Mordacia. The secretion of mucus probably protects the epithelium from the effects of digestive enzymes secreted by the zymogen cells and/or bile, which enters the diverticulum at its tip. Unlike the situation in southern hemisphere lampreys, the zymogen cells of the larvae of holarctic lampreys are located in the anterior intestine, a condition considered to be "primitive". It is thus proposed that intestinal diverticula were developed during the evolution of southern hemisphere lampreys. The relocation of zymogen cells in the diverticula increases the area for these cells, and thus the capacity for the synthesis and secretion of digestive enzymes, particularly in Mordacia where the inner surface is folded.