Fluid reabsorption by the ductuli efferentes testis of the rat is dependent on both sodium and chlorine

The role of Na(+) and Cl(-) in fluid reabsorption by the efferent ducts was examined by perfusing individual ducts in vivo with preparations of 160 mM NaCl in which the ions were replaced, together or individually, with organic solutes while maintaining the osmolality at 300 mmol/kg. Progressively replacing NaCl with mannitol reduced net reabsorption of water and the ions in a concentration-dependent manner, and caused net movement into the lumen at concentrations of NaCl less than 80 mM. The net rates of flux were lower for Na(+) than for Cl(-). In collectates, [Na(+)] was greater than [Cl(-)], indicating that Cl(-) transport is probably linked with another anion. Replacing either Na(+) or Cl(-) in perfusates (with choline and isethionate, respectively) while maintaining the other inorganic ion at 160 mM also reduced net rates of reabsorption in a concentration-dependent manner to zero when either ion was completely replaced. There were no significant differences in the osmolality of perfusate and collectate, and collectates contained a mean of 3.4 mM K(+), indicating a backflux of K(+) into the lumen. It is concluded that fluid reabsorption from the efferent ducts is dependent on the transport of both Na(+) and Cl(-) from the lumen (from a luminal concentration of at least 70-80 mM), and that Cl(-) transport is dependent on another anion. The epithelium is permeable to K(+) and has a higher permeability to a range of organic solutes (mannitol, choline, and isethionate) than epithelium in the proximal kidney tubules.     

Composition of luminal fluid secreted by the seminiferous tubules and after reabsorption by the extratesticular ducts of the Japanese quail, Coturnix coturnix japonica

The present report examines the composition of luminal fluid in the seminiferous tubule (STF), rete testis (RTF), and ductus epididymidis of the Japanese quail (Coturnix coturnix japonica). This subject is of particular interest, both because the reproductive ducts are intra-abdominal and because sperm production is more rapid in birds than in mammals. It was interpreted that micropuncture samples of STF contain varying amounts of contamination with intracellular solute, particularly K and protein. The concentration of solute in samples was correlated with packed cell volume (spermatocrit), and when the latter was used to assess estimates of solute concentration in STF, the magnitude of the estimates were much the same as determinations in RTF. Consequently, it is concluded that the fluid entering the rete testis of the quail is the primary secretion of the seminiferous tubules. The composition of RTF in the quail was determined to be 148 mM Na, 126 mM Cl, 9.8 mM K, 2.7 mM Mg, 1.4 mM Ca, 2.1 mM glutamate, 3.4 mM glutamine, 20.2 mM bicarbonate, 1.8 microg microl(-1) of protein, pH 7.34, and 310 mmol kg(-1), and it is significantly different from the composition of blood plasma. Estimates of solute output by the testis and reabsorption by the extratesticular ducts indicate, first, that most of the solutes secreted into the seminiferous tubules are subsequently reabsorbed from the extratesticular ducts and, second, that sufficient solute of testicular origin (except for protein) exists to account for the concentrations of solutes throughout the lumen of the duct system. Changes in the concentration of solute in the extratesticular ducts probably result from different reabsorption rates of solute and water. The composition of fluid from the distal end of the ductus epididymidis was 133 mM Na, 125 mM Cl, 25 mM K, 1.0 mM Mg, 0.3 mM Ca, 6.7 mM glutamate, 4.0 mM glutamine, 19.5 mM bicarbonate, 6.0 microg microl(-1) of protein, pH 7.33, and 335 mmol kg(-1), and it is significantly different from those of RTF and blood.     

New evidence for active sodium transport from fluid-filled rat lungs

The hypothesis that fluid reabsorption from the air spaces is mediated at least in part by active transport of Na+ was investigated in six sets of experiments conducted in isolated fluid-filled rat lungs. Fluid reabsorption was monitored by following the changes in the air space concentration of labeled albumin. We found that incorporation of bicarbonate rather than a nonvolatile buffer (N-2-hydroxy-ethylpiperazine-N'-2-ethanesulfonic acid) in the air space solution more than doubled the rate of fluid reabsorption. Addition of 10(-4) M amiloride to the air space solution reduced the rate of fluid reabsorption over a 2-h experiment from 1.2 +/- 0.1 to 0.7 +/- 0.1 ml and decreased reabsorption of both labeled and unlabeled Na+ from the air spaces. To show that Na+ could be reabsorbed from the air spaces even if the concentrations of Na+ in the perfusate increased above those in the air space, mannitol (150 mM) was added to the perfusate and air space solutions and the concentrations of Na+ and Cl- were reduced to 90 and 60 mM, respectively. Mannitol diffuses across the pulmonary epithelium very slowly, and it osmotically restrained the movement of water out of the air spaces. Na+ concentrations in the perfusate increased by 10 +/- 2 mM, but concentrations in the air space remained unchanged. Despite an increasingly unfavorable concentration gradient for Na+, 0.2 mmol Na+ and 0.6 ml water were reabsorbed from the air spaces in 2 h. Ouabain (10(-4) M) did not appear to slow fluid reabsorption in the presence of mannitol, but it reduced K+ secretion into the air spaces and increased K+ appearance in the perfusate in a manner consistent with inhibition of Na+-K+-adenosinetriphosphatase at the basolateral surface of the epithelial cells. Fluid reabsorption was not altered when the lungs were exposed to a hypotonic solution (185 mM), but secretion of K+ into the air spaces was accelerated and K+ was lost from the perfusate. These experiments are consistent with active Na+ transport from the air spaces.     

PGE2 enhances cytokine-elicited nitric oxide production in mouse cortical collecting duct cells.

It has been documented that arginine vasopressin (AVP) and prostaglandin E(2) (PGE(2)) regulate water reabsorption in renal tubular cells. The present study was attempted to delineate the downstream signaling of AVP and PGE(2) in a cortical collecting duct cell line (M-1 cell). Using RT-PCR, we detected mRNA for V2 and VACM-1 but not for V1a and AII/AVP receptors of AVP. Furthermore, neither AVP nor V2 receptor agonist and antagonist alter cellular cAMP. These together with unchanged cellular Ca(2+) by AVP suggested that AVP pathway was not operating in M-1 cells. All four classical PGE(2) receptors with EP3 and EP4 as the most prominent were detected in M-1 cells. PGE(2), 11-deoxy-PGE(1) (EP2 and EP4 agonist), and 17-phenyl-trinor-PGE(2) (EP1 agonist) increased cellular concentration of cAMP. There was no effect of PGE(2) or EP1 agonist on cellular Ca(2+). These findings provide evidence of the involvement of PGE(2) cascade in M-1 cells. M-1 cells were capable of synthesizing nitric oxide (NO). Although individual cytokines did not affect NO production, a mixture of tumor necrosis factor-alpha, interleukin-1beta, and interferon-gamma elevated NO concentration to 4.5-fold of the control. Addition of PGE(2) and db-cAMP to the cytokine mixture further increased NO production to 7.0- and 9.8-fold, respectively, of that seen in non-treated cells. PGE(2) or db-cAMP alone, however, had no effect on NO production. The results of the study led us to speculate that enhanced production of cAMP via PGE(2) signaling pathway in M-1 cells could either stimulate or attenuate water reabsorption in renal tubule. While an increase in cAMP alone may enhance water reabsorption, a concomitant increase in cAMP and cytokines may inhibit water reabsorption in renal tubule.     

Dibutyryl cyclic AMP decreases proliferative activity and the cholesteryl ester content in cultured cells of atherosclerotic human aorta

Dibutyryl cyclic AMP (DB cAMP) decreases by 2-5-fold the uptake of labeled thymidine into the intimal cells isolated from atherosclerotic human aorta. This effect depends on the DB cAMP concentration and the duration of its action. 0.1–1.0 mM concentration of DB cAMP reduces the level of cholesteryl esters in cells taken from atherosclerotic lesions by 1.5-2-fold. It is assumed that cAMP derivatives, and compounds which increase its intracellular content may impede the development of such atherosclerotic manifestations as smooth muscle cell proliferation and lipoidosis.     

Transport of IgG across the blood-luminal barrier of the male reproductive tract of the rat and the effect of estradiol administration on reabsorption of fluid and IgG by the epididymal ducts

In rats immunized systemically with tetanus toxoid the concentration of specific anti-tetanus-toxoid-specific IgG in fluid from the rete testis and cauda epididymidis were respectively 0.6% and 1.4% the concentration in blood serum. The extratesticular duct system reabsorbed 97% of the IgG and 99% of the fluid leaving the rete, but estradiol administration affected the site of reabsorption. In untreated rats, the ductuli efferentes reabsorbed 94% of the IgG and 96% of the fluid leaving the rete, whereas estradiol-treated rats reabsorbed 83% of the IgG and 86% of the fluid, and the ductus epididymidis fully compensated for these different effects of estradiol on the ductuli efferentes. The concentrations of IgG in secretions of the seminal vesicles and prostate gland were lower (0.1% and 0.3% respectively of the titers in blood serum) than in fluids from the extratesticular ducts, and were not affected by the administration of estradiol. RT-PCR showed that Fcgrt (neonatal Fc receptor, also known as FcRn) is expressed in the reproductive ducts, where IgG is probably transported across epithelium, being particularly strong in the ductuli efferentes (where most IgG was reabsorbed) and distal caput epididymidis. It is concluded that IgG enters the rete testis and is concentrated only 2.5-fold along the extratesticular duct system, unlike spermatozoa, which are concentrated 95-fold. Further, the ductus epididymidis can recognize and compensate for changes in function of the ductuli efferentes.     

Excretion and degradation of exogenous adenosine 3',5'-monophosphate by isolated perfused rat kidney

To determine the role played by the kidney in the metabolism and excretion of plasma adenosine 3′,5′-monophosphate (cAMP) we have studied the fate of this nucleotide (0.01–1.0mM) when it is perfused in a recirculating medium through the isolated rat kidney. cAMP was rapidly taken up and degraded by the kidney, the rate of its disappearance from the perfusate being at least twice its rate of excretion in the urine. Nevertheless, the cAMP excretory rate exceeded the filtration rate by 1.5 to 2 fold, and thus net secretion (transtubular transport) was demonstrated. The rates of filtration, perfusate clearance, and degradation of cAMP were proportional to its perfusate concentration. Methyl xanthines (caffeine and aminophylline) at 10mM, and probenecid at 0.9mM abolished transtubular transport of cAMP and greatly retarded disappearance of the nucleotide from the perfusate. It is concluded that there is a ready penetration of cAMP into renal cells from peritubular capillaries. Depending on the perfusate concentration of cAMP, transtubular transport may or may not exceed the simultaneous intra-renal breakdown of the compound. A low rate of cAMP excretion in the urine may accompany a considerably higher rate of cAMP clearance from the perfusate by the kidney.     

ATP-induced reactivation of ram testicular, cauda epididymal, and ejaculated spermatozoa extracted with Triton X-100

It was possible to demembrante and reactivate not only freshly collected testicular, cauda epididymal, and ejaculated ram sperm but also sperm that had been stored for several days at 0 degrees C and for several months at -196 degrees C in rete testis fluid or egg yolk citrate media. Sperm were usually washed free of seminal plasma before demembranation, but this was not essential for reactivation. Bovine serum albumin (1.0%) in the wash medium increased the survival of sperm, but more than 0.25% in the extraction medium decreased reactivation. A macro-molecular component of cauda epididymal fluid also inhibited the reactivation of testicular sperm. Triton X-100 concentrations between 0.01% and 1.00% in the extraction medium were satisfactory for demembranating the sperm. Rapid cooling (i.e., cold shock) mimicked the effect of detergent in making the sperm responsive to added ATP and demonstrated that damage to ram sperm in cold shock does not involve the axoneme. Ejaculated and cauda sperm were reactivated immediately on addition of ATP and activity persisted for up to 10 min. Testicular sperm, on the other hand, required about 4 min to become fully reactivated. The optimal ATP concentration for activation of sperm was 0.1-1.0 mM. Magnesium ions (0.1-1.0 mM) were important for reactivation, and testicular sperm required a higher magnesium concentration than did cauda or ejaculated sperm. Manganese ions were almost as effective as magnesium for reactivating cauda epididymal and ejaculated sperm. Cobalt and cadmium ions were much less active for cauda and ejaculated sperm and none of these ions were effective for testicular sperm. Fluoride (25-50 mM) inhibited reactivation. The presence of 50 microM cAMP in the extraction medium or preincubation of testicular sperm with theophylline or caffeine increased low levels of activation, but this was not evident with ejaculated or cauda sperm. We conclude that the motor apparatus is already functionally assembled in spermatozoa on leaving the testis, but some fine adjustment must take place during maturation in the epididymis.     

Modulation of serum-stimulated DNA synthesis in cultured human fibroblasts by cAMP

Density-dependent inhibition of growth of cultured human fibroblasts was associated with a 3- to 4-fold rise in the intracellular concentration of cyclic AMP (cAMP). Serum lowered cAMP levels in 2–5 min, with the low levels persisting for several hours. When quiescent fibroblast cultures were treated with 10% serum, the incorporation of [³H]TdR into DNA increased after a 10–16 h lag, reaching a peak by 20–24 h. Dibutyryl cyclic AMP (db-cAMP), when present throughout serum treatment, produced a dose-dependent inhibition of [³H]TdR incorporation. Half-maximal inhibition was seen with 0.1 mM db-cAMP. When db-cAMP or another cyclic nucleotide phosphodiesterase inhibitor, l-methyl-3-isobutylxanthine (SC-2964), was added together with serum to maintain elevated cAMP levels and after 4 h was replaced with fresh serum-containing medium, the wave of DNA synthesis induced by serum was not delayed. This implied that stimulation by serum could occur without an initial decrease in cAMP concentration. In contrast, db-cAMP added 8 h later than serum and not removed, inhibited [³H]TdR incorporation at the peak to the same extent as db-cAMP added together with serum. The inhibition decreased progressively when db-cAMP was added more than 8 h after serum. These results suggested that a cAMP-sensitive step occurred approx. 8 h after the addition of serum in mid-G1 of the cell cycle. Results obtained using fibroblasts synchronized at the G1/S boundary with hydroxyurea or exposed to db-cAMP for 24 h suggested that db-cAMP also inhibited TdR incorporation at the G1/S interphase or during S phase. Thus, whereas reduced cAMP concentrations did not appear to serve as an initial trigger for serum-stimulated DNA synthesis in human fibroblasts, db-cAMP and SC-2964, presumably by elevating cAMP levels, appeared to act in mid-G1 and possibly at the G1/S boundary or within S phase to inhibit thymidine incorporation.     

Fate of intraluminal glutamine in the perfused renal tubule

To determine the fate of intraluminal glutamine and specifically the role of brush border gamma glutamyltransferase in its hydrolysis and reabsorption, proximal convoluted tubules of rabbits were isolated and perfused with an artificial ultrafiltrate containing 1 mM 14C-glutamine and 3H-PEG as a volume absorption marker. The tubules, average length 0.80 +/- 0.09 mm, were bathed in perfusate containing albumin, 6.5 percent but no glutamine. Aliquots of collectate and bathing media were monitored for total 14C counts while the distribution of radioactive 14C between glutamine and glutamate in the collectate was determined by separation on a Dowex X8 formate form ion-exchange column. After 3 ten minute control periods the perfusate was switched to one containing 1 mM AT-125 in addition to glutamine and after equilibration an additional 3 collections were obtained. Control period glutamine load averaged 16.1 +/- 2.4 pmole/min of which 35 percent was absorbed and 38 and 27 percent excreted as glutamine and glutamate respectively; of the absorbed glutamine 25 percent was metabolized. During AT-125 administration, glutamine delivery averaged 15.0 +/- 2.1 pmole/min of which 57 percent was absorbed; increased absorption occurred at the expence of intraluminal glutamate formation which fell to less than 10 percent. Thus luminal transport and gamma glutamyltransferase mediated hydrolysis appear to compete for available glutamine. Significantly, reducing intraluminal glutamine hydrolysis doubles the cellular metabolism of absorbed glutamine suggesting that extracellular conversion of glutamine to glutamate alters the metabolic fate of filtered glutamine.     

In vitro inhibition of oocyte nuclear maturation in the bovine

Bovine follicular oocytes (N = 5991) were exposed to an analog of cyclic adenosine 3',5'-monophosphate (cAMP), dibutyryl cyclic AMP (db-cAMP) (2.5, 5, and 10 mM), the phosphodiesterase inhibitor isobutyl methyl xanthine (IBMX, 0.2 mM), or the purine, hypoxanthine (0.5, 1.0, 2.0 mM), and the nucleoside, adenosine (0.05, 0.1, 0.2 mM), for 6 or 21 h to assess their effects on oocyte nuclear maturation. Potential effects of bovine follicular fluid (BFF) were also evaluated after different preculture washing procedures. In a separate experiment, denuded oocytes were used to study the effect of cumulus removal on meiotic inhibition. Db-cAMP decreased the frequency of germinal vesicle breakdown (GVBD) at 6 h (88% for control and 51%, 45%, and 32% for 2.5, 5, and 10 mM concentrations, respectively). IBMX had a comparable effect with only 41% of the oocytes resuming meiosis. Hypoxanthine and adenosine alone or in combinations also decreased the number of oocytes undergoing GVBD at 6 h. Only 22% GVBD occurred when the combined highest concentration of both substances was used compared to 88% in controls. If oocytes were incubated in 50% BFF after a wash in control medium during processing, 56% would resume meiosis at 6 h vs. 35% if the washing procedure included inhibitors (db-cAMP + IBMX). Total BFF (100%) during washing and maturation prevented 72% of the oocytes from resuming meiosis. Db-cAMP and IMBX combined or BFF also inhibited meiotic resumption of denuded oocytes. At 21 h, the inhibitory effects were less pronounced, with most oocytes only delayed in completing the first reduction division.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of dibutyryl cyclic AMP and isoproterenol on 7 beta-hydroxycholesterol cytotoxicity and esterification in spontaneous transformed cell lines derived from astrocyte primary cultures.

Incubation of spontaneous transformed cells derived from astrocyte primary cultures with 30 microM 7 beta-hydroxycholesterol (7 beta-OH-CH) which is lethal to the cells or with 150 microM isoproterenol reduces the intracellular level of cAMP (4- and 2-fold respectively). Treatment of the cultures with 0.5 mM dibutyryl (db)-cAMP and 7 beta-OH-CH increases 3-fold the intracellular level of cAMP and both, db-cAMP and isoproterenol, raise the lethal effect of 7 beta-OH-CH and its esterification on C-3-OH by naturally occurring fatty acids (metabolite). Kinetic studies of net steryl-3-esters hydrolysis revealed that db-cAMP and isoproterenol lower that of cholesteryl-3-esters (2-fold) whereas the opposite is found for the metabolite. These data demonstrate that (i) high cAMP intracellular levels modulate differently the net hydrolysis of cholesteryl-3-esters and metabolite, (ii) isoproterenol acts otherwise than cAMP on 7 beta-OH-CH esterification, (iii) the cytotoxicity of 7 beta-OH-CH is linked to its own esterification. The accumulation of metabolite subsequent to db-cAMP or isoproterenol treatment as a result of acyl-CoA:cholesterol acyl transferase activation is discussed.     

Pleural surface fluorescence measurement of Na+ and Cl- transport across the air space-capillary barrier.

We developed a pleural surface fluorescence method to measure Na(+) and Cl(-) transport in perfused mouse lungs. The air space was filled with aqueous fluid containing membrane-impermeant fluorescent indicators of Cl(-) (lucigenin) or Na(+) (Sodium Green). After instillation of a Cl(-)-free solution into the air space, an increase in perfusate Cl(-) concentration from 0 to 30 mM produced a decrease in surface lucigenin fluorescence (6.5%/min) corresponding to Cl(-) influx of 1.0 mM/min. Cl(-) influx was increased to 2.1 +/- 0.3 mM/min by forskolin, and the increase was inhibited by glibenclamide. cAMP-stimulated Cl(-) influx was decreased by 57% in CFTR null mice. After instillation of a Na(+)-free solution into the air space, an increase in perfusate Na(+) concentration from 0 to 30 mM gave increased Sodium Green fluorescence (Na(+) influx of 1.2 mM/min), which increased approximately fivefold after cAMP agonists. Cl(-) and Na(+) transport were not affected in lungs from mice lacking aquaporins AQP1 or AQP5. Our results establish a pleural surface fluorescence method to measure unidirectional Cl(-) and Na(+) flux in intact lung and provide evidence for cAMP-stimulated transcellular Cl(-) and Na(+) transport.     

Biphasic effect of calcium on luteinizing hormone-stimulated cyclic adenosine 3',5'-monophosphate production in granulosa cells of the fowl (Gallus domesticus).

Both cAMP and Ca2+ play important roles in the steroidogenic action of LH in hen granulosa cells. However, the interaction of these intracellular messengers is not fully understood. In the present study we used two calcium ionophores (ionomycin and A23187), as well as trifluoperazine (TFP), an inhibitor of calmodulin, to investigate LH- and forskolin-induced cAMP production in granulosa cells isolated from the largest (F1) preovulatory follicle of White Leghorn laying hens. Between 0.1 and 1.0 microM, both ionophores significantly potentiated cAMP responses to LH in the presence of 0.1 mM extracellular Ca2+. When calcium was omitted from the medium, ionophores had no effect. When either calcium was raised above 1 mM, or the concentration of ionophores was increased above 1 microM, LH-induced cAMP production was drastically inhibited. In the presence of 0.5-2.0 mM calcium, A23187 inhibited forskolin-promoted cAMP synthesis. TFP, while having no effect on basal cAMP, suppressed LH-induced responses and the potentiating effect of ionomycin. It is concluded that for full activation of the adenylate cyclase/cAMP system by LH, Ca-calmodulin is required at a site upstream from the catalytic component of the enzyme. However, high intracellular Ca2+ and/or other effects of ionophores (such as uncoupling of oxidative phosphorylation) inhibit LH-induced cAMP production.     

Metabolism of valine and 3-methyl-2-oxobutanoate by the isolated perfused rat kidney.

Metabolism of branched-chain amino and 2-oxo acids was studied in the isolated perfused kidney. Significant amounts of 2-oxo acids were released by perfused kidney with all concentrations of amino acids tested (0.1-1.0 mM each), despite the high activity of branched-chain 2-oxo acid dehydrogenase in kidney. As perfusate valine concentration was increased from 0.2 to 1.0 mM, [1-14C]valine transamination (2-oxo acid oxidized + released) increased roughly linearly; [1-14C]valine oxidation, however, increased exponentially. Increasing perfusate concentration of 3-methyl-2-oxo[1-14C]butanoate from 0 to 1.0 mM resulted in a linear increase in the rate of its oxidation and a rise in perfusate valine concentration; at the same time significant decreases occurred in perfusate isoleucine and leucine concentrations, with corresponding increases in rates of release of their respective 2-oxo acids. Comparison of rates of oxidation of [1-14C]valine and 3-methyl-2-oxo[1-14C]butanoate suggests that 2-oxo acid arising from [1-14C]valine transamination has freer access to the 2-oxo acid dehydrogenase than has the 2-oxo acid from the perfusate. The observations indicate that, when branched-chain amino and 2-oxo acids are present in perfusate at near-physiological concentrations, rates of transamination of the amino and 2-oxo acids by isolated perfused kidney are greater than rates of oxidation.     

The role of cyclic nucleotides in the regulation of mitotic activity in SSPE virus-infected human brain tissue.

The intracellular level of cGMP was independent of the rate of cell division in cells derived from virally infected brain tissue. The phosphodiesterase inhibitor R07-2956 (4-dimethoxybenzyl-2-imidazolidinone) increased the intracellular level of cGMP in virally infected brain cells, but it did not effect the level of cAMP. There was no correction between the increase in cGMP levels following addition of R07-2956 and changes in mitotic activity in the brain cell cultures. Experimental manipulations which increased the cAMP level were accompanied by a decreased mitotic rate indicating there was a correlation between mitotic activity and the level of cAMP in the same cells. Raising the intracellular level of cAMP by exogenous db-cAMP or cAMP or the use of other phosphodiesterase inhibitors routinely increased the level of cGMP as well. Conversely increasing the intracellular cGMP level by adding the exogenous cGMP increased the level of both cGMP and cAMP.A tissue culture system was used with the cell line derived from viral infected human brain tissue originally obtained from a patient with subacute sclerosing panencephalitis (SSPE). The intracellular levels of cAMP and cGMP were monitored by radioimmunoassay following manipulation of the system by addition of exogenous cGMP (0.05 mM), addition of exogenous db-cAMP (0.5 mM), or cAMP (0.5 mM) and the use of phosphodiesterase inhibitors: theophylline (1.0 mM), papaverine (50 μg/ml), 4-3-butoxy-4-methoxy benzyl-2-imidozalidinone (R020-1724) and R07-2956. Cell division was monitored in treated and non-treated cultures at 24 h intervals by analyzing the cell number and mitotic index.High levels of cGMP were found in cells which were not actively dividing but high levels were just as apt to be present in dividing cells. There was an inverse relationship between cell division and the level of cAMP.     

Effects of L-glucose on sodium reabsorption in the isolated perfused rat kidney.

In the isolated perfused rat kidney, sodium reabsorption is enhanced in the presence of 5.5 mM D-glucose. However, it is unclear whether this effect is metabolic or whether it is due to a requirement for sodium transport in the process of glucose reabsorption. A third possibility is solvent drag. In an attempt to differentiate between these possibilities, kidneys were perfused with the D-glucose isomer, L-glucose (L-G), a nonmetabolizable hexose. At a perfusate concentration of 5.5 mM L-G, per cent L-G reabsorption was approximately 30. Inhibition of L-G reabsorption by D-glucose suggests carrier-mediated transport. In the presence of 5.5 mM L-G, sodium reabsorption approximated 92% during the course of perfusion. When L-G was omitted from perfusate, sodium reabsorption ultimately declined to 85%. Since significant metabolism of L-G was not observed, the results are compatible with the hypothesis that enhanced sodium reabsorption may be brought about by some still to be defined aspect of glucose transport.     

Differential effects of various cAMP derivatives on the morphological and electrical maturation of neuroblastoma X glioma hybrid cells

The influence of adenosine 3′: 5′-cyclic monophosphate (cAMP) and some of its derivatives on the morphological differentiation and on the expression of electrical activity was investigated in neuroblastoma X glioma hybrid cells. This permanent cell line constitutes a well established culture system for studying neuronal properties in vitro. cAMP (1 mM) caused cell death. With 8-bromo-cAMP (0.1–1 mM) present giant multinuclear cells appeared, which were more obvious at 0.1 than at 1 mM 8-bromo-cAMP. 8-p-chlorophenylthio-cAMP (0.1–1 mM) induced an extension of neurites. These cellular processes were comparable to those elicited in the presence of db-cAMP (1 mM). However, only the cells treated with db-cAMP, but not those exposed to 8-p-chlorophenylthio-cAMP, were found to generate action potentials upon electrical stimulation. Neither dexamethasone nor carboxymethylcellulose, nor 8-bromo-cAMP could elicit the formation of processes in hybrid cells.     

Amiloride inhibits contraction and serotonin modulation of Aplysia muscle

1. Serotonin (5-HT) potentiates acetylcholine (ACh)-elicited contractions of Aplysia buccal muscles. Serotonin potentiation was significantly reduced by 0.03 mM, 0.1 mM, and 0.3 mM amiloride. 2. Unpotentiated ACh-elicited contractions were significantly reduced by 0.1 mM and 0.3 mM amiloride. 3. Amiloride reduced ACh-elicited depolarization. The reduction in contraction caused by 0.3 mM amiloride (to 16% of control) was larger than could be explained by the reduction in depolarization (86% of control). 4. Amiloride had no effect on tension in skinned muscle fibers, indicating that amiloride probably did not have a direct effect on contractile mechanisms. 5. Potentiation of contraction produced by zero sodium (Tris substituted, 0 Na-Tris) medium could be abolished by 0.3 mM amiloride. 6. Zero Na-Tris increased 45Ca influx 2.7-fold. In the presence of 0.3 mM amiloride, 0 Na-Tris increased 45Ca influx only 1.4-fold. 7. Amiloride (0.3 mM) reduced the elevation of muscle cAMP caused by 10(-6) M 5-HT by 60%. Zero Na-Tris did not cause a change in muscle cAMP.     

Human müllerian inhibitory factor messenger ribonucleic acid is hormonally regulated in the fetal testis and in adult granulosa cells

The regression of the Müllerian ducts (the embryologic precursor of uterus, vagina, and Fallopian tubes) in the male fetus is caused by Müllerian inhibitory factor (MIF), a glycoprotein produced by fetal Sertoli cells. Although this Müllerian duct involution is complete before midgestation, the amount of MIF mRNA did not vary among 25 human fetal testis samples from 13 to 25.8 weeks of gestation. In cultured 20-week human testis cells, cAMP increased MIF mRNA 8.3-fold, but the human gonadotropins FSH and CG had no effect. In cultured adult human granulosa cells, CG and cAMP increased MIF mRNA accumulation to 430% and 890%, respectively, but FSH had no effect. The expression and hormonal regulation of MIF mRNA in midgestation testes and in adult granulosa cells indicate that MIF has physiological roles in the human gonad other than Müllerian duct regression.