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The CFTR High Expresser (CHE) cells express eightfold higher levels of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl−channel compared with neighboring enterocytes and were first identified by our laboratory (Ameen et al., Gastroenterology 108: 1016, 1995). We used double-label immunofluorescence microscopy to further study these enigmatic epithelial cells in rat intestine in vivo or ex vivo. CHE cells were found in duodenum, most frequent in proximal jejunum, and absent in ileum and colon. CFTR abundance increased in CHE cells along the crypt-villus axis. The basolateral Na+K+Cl−cotransporter NKCC1, a key transporter involved in Cl−secretion, was detected at similar levels in CHE cells and neighboring enterocytes at steady state. Microvilli appeared shorter in CHE cells, with low levels of Myosin 1a, a villus enterocyte-specific motor that retains sucrase/isomaltase in the brush-border membrane (BBM). CHE cells lacked alkaline phosphatase and absorptive villus enterocyte BBM proteins, including Na+H+exchanger NHE3, Cl−/HCO3−exchanger SLC26A6 (putative anion exchanger 1), and sucrase/isomaltase. High levels of the vacuolar-ATPase proton pump were observed in the apical domain of CHE cells. Levels of the NHE regulatory factor NHERF1, Na-K-ATPase, and Syntaxin 3 were similar to that of neighboring enterocytes. cAMP or acetylcholine stimulation robustly increased apical CFTR and basolateral NKCC1 disproportionately in CHE cells relative to neighboring enterocytes. These data strongly argue for a specialized role of CHE cells in Cl−-mediated “high-volume” fluid secretion on the villi of the proximal small intestine.

Changes in intestinal luminal pH affect mucosal ion transport. The aim of this study was to compare how luminal pH and specific second messengers modulate the membrane traffic of four major ion transporters (CFTR, NHE3, NKCC1, and NBCe1) in rat small intestine. Ligated duodenal, jejunal, and ileal segments were infused with acidic or alkaline saline, 8-Br-cAMP, or the calcium agonist carbachol in vivo for 20 min. Compared with untreated intestine, lumen pH was reduced after cAMP or carbachol and increased following HCO3−-saline. Following HCl-saline, lumen pH was restored to control pH levels. All four secretory stimuli resulted in brush-border membrane (BBM) recruitment of CFTR in crypts and villi. In villus enterocytes, CFTR recruitment was coincident with internalization of BBM NHE3 and basolateral membrane recruitment of the bicarbonate transporter NBCe1. Both cAMP and carbachol recruited NKCC1 to the basolateral membrane of enterocytes, while luminal acid or HCO3−retained NKCC1 in intracellular vesicles. Luminal acid resulted in robust recruitment of CFTR and NBCe1 to their respective enterocyte membrane domains in the upper third of the villi; luminal HCO3−induced similar membrane changes lower in the villi. These findings indicate that each stimulus promotes a specific transporter trafficking response along the crypt-villus axis. This is the first demonstration that physiologically relevant secretory stimuli exert their actions in villus enterocytes by membrane recruitment of CFTR and NBCe1 in tandem with NHE3 internalization.

We examined the cell-specific subcellular expression patterns for sodium- and potassium-coupled chloride (NaK2Cl) cotransporter 1 (NKCC1), Na+bicarbonate cotransporter (NBCe1), cystic fibrosis transmembrane conductance regulator (CFTR), and Na+/H+exchanger 3 (NHE3) to understand the functional plasticity and synchronization of ion transport functions along the crypt-villus axis and its relevance to intestinal disease. In the unstimulated intestine, all small intestinal villus enterocytes coexpressed apical CFTR and NHE3, basolateral NBCe1, and mostly intracellular NKCC1. All (crypt and villus) goblet cells strongly expressed basolateral NKCC1 (at approximately three-fold higher levels than villus enterocytes), but no CFTR, NBCe1, or NHE3. Lower crypt cells coexpressed apical CFTR and basolateral NKCC1, but no NHE3 or NBCe1 (except NBCe1-expressing proximal colonic crypts). CFTR, NBCe1, and NKCC1 colocalized with markers of early and recycling endosomes, implicating endocytic recycling in cell-specific anion transport. Brunner's glands of the proximal duodenum coexpressed high levels of apical/subapical CFTR and basolateral NKCC1, but very low levels of NBCe1, consistent with secretion of Cl−-enriched fluid into the crypt. The cholinergic agonist carbachol rapidly (within 10 min) reduced cell volume along the entire crypt/villus axis and promoted NHE3 internalization into early endosomes. In contrast, carbachol induced membrane recruitment of NKCC1 and CFTR in all crypt and villus enterocytes, NKCC1 in all goblet cells, and NBCe1 in all villus enterocytes. These observations support regulated vesicle traffic in Cl−secretion by goblet cells and Cl−and HCO3−secretion by villus enterocytes during the transient phase of cholinergic stimulation. Overall, the carbachol-induced membrane trafficking profile of the four ion transporters supports functional plasticity of the small intestinal villus epithelium that enables it to conduct both absorptive and secretory functions.

We previously demonstrated greater HPA axis activation in adult men compared to adult women following low-dose administration of the anticholinesterase inhibitor, physostigmine (PHYSO). Because blood sampling was done infrequently following PHYSO, the rise times of AVP, ACTH1-39, and cortisol could not be determined. In the present study, we determined the sequence of hormone increases by frequent blood sampling following PHYSO. Twelve adult women and 12 adult men underwent three test sessions 5-7 days apart: PHYSO, saline control, and repeat PHYSO. As in the earlier study, PHYSO produced no side effects in half the subjects and mild side effects in the other half, with no significant female-male differences. None of the hormone responses was significantly correlated with the presence or absence of side effects. In both women and men, the AVP increase preceded the ACTH1-39 increase, which in turn preceded the cortisol increase. The AVP and ACTH AUCs were significantly positively correlated in both women and men, supporting AVP as an acute stimulus to ACTH secretion. Also as in the earlier study, the AVP response to PHYSO was more than twice as great in men as in women, but the difference was not statistically significant. We therefore analyzed the results of both studies combined (N=26 women and 26 men). The men had a significantly greater AVP response and a trend toward a greater ACTH1-39 response compared to the women. These findings further support the concept of sexual diergism (functional sex difference) in the influence of CNS cholinergic systems on HPA hormone secretion.

The transition from the wood-and-fabric airplane to the all-metal airplane was essentially complete by World War II. The late 1920s and early 1930s are said to have witnessed a structural revolution in aeronautics with the appearance of streamlined metal aircraft with such features as tightly cowled multiple engines, variable-pitch propellers, retracting landing gear, and stressed-skin aluminum construction. A prevalent assumption regarding this transition is that the building material acted as a primary driver of change, that engineering advance was guided by an inevitable move toward metal structures. Metal did indeed allow engineers to extend performance parameters afforded by innovative structural designs, but, interestingly, many of these key innovations were not developed to takeadvantageof metal. They emerged independent of theconstruction material, and often weree rst used in wooden airplanes. The cluster of original ideas that coalesced in the 1930s constituted one of the major watershedperiodsin aerospacetechnology.Metalcarriedthisbasicdesign revolutiontothelimitsofitsengineering andtechnicalfeasibility,butonlyafteranewfoundationwasinplace.Metaldidnotspawnthestructuralrevolution. An exploration of therootsof the structural revolution in aeronautics and thecomplexity oftechnological progress is presented.

Reorganization of the cerebellar glomerulus, the main synaptic complex within the granule cell layer, was investigated using quantitative morphological techniques. All afferents to the cerebellar cortex, including mossy-fibers, were surgically destroyed by undercutting the cerebellar vermis. Fifteen days after the operation, which resulted in the removal of the main excitatory afferent to the glomerulus, a significant reorganization of the whole synaptic complex was observed, whereas the structural integrity of the glomerulus was remarkably well preserved. This was indicated by the observation that the number of granule cell dendrites (approximately 50 per glomerulus), as well as the number of dendritic digits (approximately 210 per glomerulus) bearing most of the approximately 230 synaptic junctions per glomerulus, did not change significantly after mossy-fiber degeneration. The total number of synapses in the reorganized glomerulus did not change either, despite the disappearance of two-thirds of (excitatory) synaptic junctions caused by mossy-fiber degeneration. In the reorganized glomeruli, however, the inhibitory, GABA-containing Golgi axonal varicosities became the dominant synaptic type-about four-fifths (approximately 200) of all synapses within the glomerulus-whereas the dendritic synapses between the granule cells represented only one-fifth of all synaptic junctions. The quantitative data of the reorganized cerebellar glomerulus demonstrate both a remarkable constancy and a plasticity of the excitatory granule cells and inhibitory Golgi neurons building up this synaptic complex. Constancy (the preservation of certain specific structural features) is represented by an eventually unchanged number of dendrites and synaptic junctions within the deafferented glomerulus. Such constancy was made possible, however, by the morphogenetic plasticity of both nerve-cell types to produce new, dendro-dendritic and axo-dendritic synapses to compensate for the loss of mossy-fiber synapses.

Intraneuronal production of estradiol from testosterone has been shown to play a pivotal role in gender-specific brain development of most vertebrates, and to participate in numerous functions of the adult central nervous system. Previous biochemical and morphological approaches demonstrated that estrogen synthetase (aromatase) is present in specific limbic and hypothalamic structures. On the other hand, less attention has been paid to revealing its subcellular distribution. The possibility of aromatase presence in axonal processes has been indicated by recent biochemical and morphological observations suggesting new insights for the role of aromatase in neural functions. The objective of the present study was to provide morphological evidence for the subcellular location of aromatase in neurons of different vertebrate species including Japanese quail, rat, monkey, and human. Immunocytochemistry using a purified polyclonal antiserum against human placental aromatase localized immunoreactivity to hypothalamic and limbic cell groups in all of these species. Light and electron microscopic examination of vibratome sections revealed the presence of aromatase immunoreactivity throughout the neuronal perikarya, including dendrites and axonal processes. In each species there were numerous boutons which contained labeled small clear synaptic vesicles. Many of these axon terminals formed synapses with immuno-negative and immuno-positive dendrites and perikarya. This study furnishes the first immunolocalization of aromatase in the brains of two primate species, humans and monkeys. The provision of further evidence for estrogen synthesis in axons and axon terminals may help resolve apparent differences between the measurement of aromatase activity and the lack of aromatase-immunopositive cell bodies in previous studies. The present findings may be coupled with recent evidence regarding the molecular biology and the diversity of functional properties of P450 aromatase to indicate previously unexpected effects of brain aromatase at the synaptic level.

We observed significantly reduced serum alpha 2-HS glycoprotein concentrations in patients with acute lymphocytic, acute nonlymphocytic, chronic granulocytic and chronic myelomonocytic leukemias, Hodgkin's and non-Hodgkin's lymphomas, myelofibrosis, and multiple myeloma, but not in patients with chronic lymphocytic leukemia and polycythemia vera, as compared with healthy controls. We followed the serum level of the protein for 18 months. Patients with infectious complications, those receiving cytostatic treatment, and those in the preterminal period had further reduced serum alpha 2-HS glycoprotein levels. The reduction of serum alpha 2-HS glycoprotein concentration was primarily due to decreased production caused by infiltration of the liver, a hepatotoxic effect of cytostatic treatment, and, to a lesser degree, to increased consumption. We found statistically significant negative correlations between serum alpha 2-HS glycoprotein concentration and erythrocyte sedimentation rate, serum aspartate aminotransferase and alkaline phosphatase activities, and IgG and IgM concentrations. The determination of the alpha 2-HS glycoprotein concentration is useful for the assessment and follow-up of the clinical status and therapy of patients with hematological malignancies and also has prognostic significance.

Soluble L-selectin (sL-selectin) concentrations were measured in paired samples of serum and cerebrospinal fluid by an ELISA method. Patients with several forms of multiple sclerosis (MS) and systemic lupus erythematosus with central nervous system involvement (SLE-CNS) were investigated. Elevated CSF sL-selectin concentrations were found in patients with SLE-CNS (7.62 +/- 3.31 ng/ml) and with relapsing-remitting form of MS (6.99 +/- 4.72 ng/ml) compared to the control group (4.00 +/- 0.95 ng/ml). The data presented suggest some similarities between inflammatory/immunological events in the central nervous system in patients with SLE-CNS and relapsing-remitting form of MS. Immunological heterogeneity in MS is suspected.

Interleukin (IL) 6 has an important role in the regulation of acute-phase proteins (APPs) during an acute-phase response. We studied IL-6 and other cytokines to determine if they regulate serum APP levels in the same way under the condition of the aberrant, long-lasting 'acute-phase response' that occurs in patients with chronic inflammation and cancer.Serum levels of nine positive APPs [CRP, SAA, C1-INH, Bf, C5, C8, C9, alpha 1-acidic glycoprotein (AGP) and haptoglobin] and two negative APPs [transferrin and alpha 2-HS glycoprotein (AHSG)] were measured using immunochemical methods in 59 multiple myeloma patients and in 72 healthy control subjects. Serum IL-6 and tumour necrosis factor (TNF) alpha levels were determined by bioassays.IL-6 was negatively correlated with five out of nine (C1-INH, C8, C9, AGP and haptoglobin) positive APPs but positively correlated with C-reactive protein (CRP). When patients with high and low IL-6 serum concentration were compared, CRP levels were higher, AGP and haptoglobin levels were lower in the high- than in the low-L-6 group, whereas no significant difference between the two groups was found in levels of the other positive and negative APPs. TNF-alpha levels were negatively correlated with transferrin and AHSG levels. No difference in the levels of positive APPs was observed between patients with low and high TNF-alpha serum concentration. By contrast, levels of both transferrin and AHSG were significantly lower in the high- than in the low-TNF-alpha group.These findings indicate that, except for regulation of the negative APPs by TNF-alpha, the mechanism of APP regulation is different under the conditions of the short-term and the chronic, long-lasting 'acute-phase reaction'.

The mediolateral part of the lateral septal area (LSA) is a common target of hippocamposeptal afferents, neuropeptide containing, catecholaminergic, cholinergic, and GABAergic pericellular baskets of different origins. This specific innervation pattern as well as electrophysiological data concerning this area suggest a convergent input from different sources to particular LSA neuron populations. Light and electron microscopy combined with Golgi impregnation and acute anterograde degeneration techniques following transection of the fimbria-fornix were employed to determine whether LSA neurons with hippocampal input have any characteristic and distinctive morphological signs. About 20% of all Golgi impregnated LSA neurons were found to have somatic spines. All of these somatospiny neurons are synaptic targets of hippocamposeptal fibers. The degenerated hippocamposeptal boutons establish asymmetric synaptic contacts on their soma, somatic and dendritic spines, and on dendritic shafts. Somatospiny neurons located in the most medial and dorsal parts of the LSA seem to project toward the medial septum while all of the others appear to send descending fibers to ventral areas. Somatospiny neuron axons occasionally give out recurrent collaterals. Quantitative analysis on the spatial distribution of the somatospiny neurons revealed that practically all of them are encountered in the mediolateral division of the LSA. This area includes the lateral part of the intermediolateral septal nucleus and adjacent lateral portions of the dorsolateral and the ventrolateral septal nuclei.

Computer-assisted stereological and quantitative morphological approaches were used to analyse cerebellar glomeruli of the “simple type” in serial ultrathin sections. It was found that, of the total volume (110–200 μm3) of the glomeruli studied, 53% was occupied by granule cell dendrites, 34% by mossy terminal and 13% by Golgi axons. None of the four analysed glomeruli contained Golgi cell dendrites. The mossy terminals that were studied received, on the average, 53 granule cell dendrites. All of the dendrites originated from different granule cells and all made synaptic contacts with mossy terminal. However only about 60% of granule cell dendrites made synapses with Golgi axons. The surface of the mossy terminals occupied by synaptic junctions, was found to be 5.4–5.5%. Each granule cell dendrite emitted 3–5 terminal protrusions (“dendritic digits”). Each digit receives one or more synaptic contact from either the mossy terminal (67% of all digits), or from Golgi axon varicosities (25%). Only about 8% of all digits were contacted synaptically by both types of axonal terminals. All of the dendritic digits that were observed made synaptic connections. Each digit was, on the average, connected by symmetric attachment plaques to 4 neighbouring digits. Three-dimensional reconstructions of mossy terminal and some of contacting granule cell dendrites demonstrated that the dendrites curved around the central mossy terminal and were much longer than expected from earlier Golgi-impregnation studies. In addition to mossy terminals and Golgi axons, an axon terminal of small calibre that contained large, empty, spheroid vesicles were occasionally observed. These terminals, which are most likely the axonal varicosities of ascending parallel fibers, made synaptic contacts exclusively with granule cell dendrites at the periphery of the glomeruli. The results demonstrate that, in the rat cerebellum, there is a high degree of convergence of granule cells at a glomernius (53 to 1); and that there is a rich inhibitory input to about 60% of all granule cell dendrites. It is also shown that the main postsynaptic targets, for both mossy and Golgi axons, are the dendritic digits. The presence of synaptic contacts between parallel-fiber-like varicosities and granule cell dendrites may be an additional source of excitation within the glomerulus.

There is a diminution of D-glucose absorption in the aged rat which is partly due to the decrease of the length of the villi.