Erythrocyte Na+, K+-ATPase activity in patients with congestive heart failure.

In order to determine if the Na+, K+-ATPase activity in erythrocyte membranes is altered in congestive heart failure, and to examine its clinical significance with respect to other clinical variables, erythrocyte Na+, K+-ATPase activity was measured in 51 patients with left ventricular ejection fractions <40% (coronary artery disease, n=26; dilated cardiomyopathy, n=25) and 24 control patients. Na+, K+-ATPase activity was lower in both coronary artery disease and dilated cardiomyopathy groups than control group even in the absence of digitalis use. There was a significant inverse correlation between Na+, K+-ATPase activity and plasma norepinephrine. The presence of non-sustained ventricular tachycardia was associated with a lower Na+, K+-ATPase activity in both groups with congestive heart failure without digitalis use than those without ventricular tachycardia. Plasma norepinephrine was higher in patients with non-sustained ventricular tachycardia than those without in the coronary artery disease group, but not in the dilated cardiomyopathy group. Na+, K+-ATPase activity may be helpful in predicting electrophysiologic instability in patients with heart failure.     

cAMP regulates nitric oxide production and ouabain sensitive Na+, K+-ATPase activity in SH-SY5Y human neuroblastoma cells

Summary We investigated the relation between cyclic AMP (cAMP) and nitric oxide (NO) production, as well as the effect of NO on Na⁺ , K⁺-ATPase activity in the human neuroblastoma cell line SH-SY5Y. Two cAMP agonists, dibutyryl cAMP (DBC) and beraprost sodium (BPS), increased cAMP accumulation and NO production in a time and dose dependent manner at 50 mmol/l glucose. On the other hand, cellular sorbitol and myo-inositol contents and protein kinase C activity were not altered by DBC or BPS. A specific protein kinase A inhibitor, H-89, suppressed increases in nitrite/nitrate and cyclic GMP (cGMP) and protein kinase A activity stimulated by DBC or BPS. This finding suggests that cAMP stimulates NO production by activating protein kinase A via a pathway different from the sorbitol-myo-inositol-protein kinase C pathway. We observed that an NO donor, sodium nitroprusside, and an NO agonist, L-arginine, enhanced ouabain sensitive Na⁺, K⁺-ATPase activity at 50 mmol/l glucose. We also found that a nitric oxide synthase inhibitor, N^G-nitro-L-arginine methyl ester (L-NAME), inhibited Na⁺, K⁺-ATPase activity at 5 mmol/l glucose, and partially suppressed the enzyme activity stimulated by DBC or BPS. The results of this study suggest that cAMP regulates protein kinase A activity, NO production and ouabain sensitive Na⁺, K⁺-ATPase activity in a cascade fashion. The results also suggest that protein kinase A at least partially regulates Na⁺, K⁺-ATPase activity without mediation by NO in SH-SY5Y cells. We speculate that cAMP and NO are two important regulatory factors in the pathogenesis of diabetic neuropathy. [Diabetologia (1998) 41: 1451–1458] Received: 17 November 1997 and in final revised form: 27 July 1998     

The effect of thyroxine on (Na+, K+)-ATPase from the heart and the kidney of rabbit.

The effect of thyroxine on membrane bound (Na+, K+)-ATPase isolated as a microsomal fraction from rabbit heart and kidney was investigated. In the heart, thyroxine administration produced an increased Ki value (a concentration of ouabain required for half maximal inhibition of (Na+, K+)-ATPase activity) without alteration of the specific activity of cardiac (Na+, K+)-ATPase, indicating that the digitalis sensitivity of cardiac (Na+, K+)-ATPase was decreased. On the contrary, a significant increase of the specific activity of renal (Na+, K+)-ATPase was observed without change in its digitalis sensitivity. These results suggest that (1) a decreased sensitivity of cardiac (Na+, K+)-ATPase to digitalis glycosides in thyrotoxic animals may contribute to the decrease in the inotropic and toxic effects of the digitalis glycosides in the hyperthyroid state, and that (2) there may be an organ difference in (Na+, K+)-ATPase.     

The cyclic AMP/protein kinase A signal transduction pathway modulates tolerance to sedative and hypothermic effects of ethanol

Background: An expanding body of literature indicates the important role of the cAMP/PKA signaling pathway in establishing initial sensitivity to alcohol as well as being involved in certain forms of tolerance to ethanol. The use of mice with heterozygous inactivation of the Gnas gene encoding Gsα allowed us to explore the relationship between tolerance to ethanol and cAMP/PKA signaling. Methods: Mice with the targeted disruption of one Gsα allele were compared with wild‐type littermates in their initial sensitivity to ethanol‐induced sedation and hypothermia and then monitored for the development of tolerance during two subsequent bouts of intoxication. Components of the cAMP/PKA signaling pathway were analyzed in ethanol‐naïve mice and again following the development of tolerance to ethanol to better understand the contribution of this signaling pathway to the acquisition of tolerance. Results: During the initial exposure to ethanol, mice with the targeted disruption of one Gsα allele (Gnas) were more sensitive to the sedative effects of ethanol compared with wild‐type littermates. Wild‐type mice developed within‐session tolerance to ethanol‐induced hypothermia whereas Gnas mice did not. Following the subsequent ethanol treatments, wild‐type mice developed between‐session tolerance to the sedative effects of ethanol to a greater degree than mice with heterozygous inactivation of the Gnas gene. The development of tolerance to the sedative effects of ethanol was accompanied by increased expression of phospho‐CREB in the cerebellum, hippocampus, and frontal cortex. No changes in phospho‐CREB expression were detected in these brain regions in mice with heterozygous inactivation of the Gnas gene. Conclusion: The results show that cAMP/PKA signal transduction modulates sensitivity to sedative and hypothermic effects of ethanol. This signal transduction system also influences the acquisition of within‐session and between‐session tolerance. The mechanism through which cAMP/PKA signaling modulates the development of tolerance remains to be elucidated but may involve changes in phospho‐CREB expression.     

Protein kinase C effects on nerve function, perfusion, Na+,K+-ATPase activity and glutathione content in diabetic rats

Abstract Aims//hypothesis. Increased protein kinase C activity has been linked to diabetic vascular complications in the retina and kidney, which were attenuated by protein kinase C antagonist treatment. Neuropathy has a vascular component, therefore, the aim was to assess whether treatment with WAY151 003 or chelerythrine, inhibitors of protein kinase C regulatory and catalytic domains respectively, could correct nerve blood flow, conduction velocity, Na⁺,K⁺-ATPase, and glutathione deficits in diabetic rats. Methods. Diabetes was induced by streptozotocin. Sciatic nerve conduction velocity was measured in vivo and sciatic endoneurial perfusion was monitored by microelectrode polarography and hydrogen clearance. Glutathione content and Na⁺,K⁺-ATPase activity were measured in extracts from homogenised sciatic nerves. Results. After 8 weeks of diabetes, sciatic blood flow was 50 % reduced. Two weeks of WAY151 003 (3 or 100 mg/kg) treatment completely corrected this deficit and chelerythrine dose-dependently improved nerve perfusion. The inhibitors dose-dependently corrected a 20 % diabetic motor conduction deficit, however, at high doses ( > 3.0 mg/kg WAY151003; > 0.1 mg/kg chelerythrine) conduction velocity was reduced towards the diabetic level. Sciatic Na⁺,K⁺-ATPase activity, 42 % reduced by diabetes, was partially corrected by low but not high dose WAY151 003. In contrast, only a very high dose of chelerythrine partially restored Na⁺,K⁺-ATPase activity. A 30 % diabetic deficit in sciatic glutathione content was unchanged by protein kinase C inhibition. The benefits of WAY151 003 on blood flow and conduction velocity were blocked by nitric oxide synthase inhibitor co-treatment. Conclusion/interpretation. Protein kinase C contributes to experimental diabetic neuropathy by a neurovascular mechanism rather than through Na⁺,K⁺-ATPase defects. [Diabetologia (1999) 42: 1120–1130] Received: 21 December 1998 and in final revised form: 23 April 1999     

Reduced Na+, K+ -ATPase activity in intact red cells and isolated membranes from obese man

Na+, K+ -ATPase activity was measured in red blood cells from 20 nondiabetic euthyroid male Pima Indians with varying degrees of obesity; their body mass indices ranged from 22-60 kg/m2. The na+, K+ -ATPase, measured both by 86Rb uptake in intact cells and ATP hydrolysis by purified membranes, was inversely correlated with body mass index (r = -0.62; P less than 0.005 and r = -0.75; P less than 0.0001, respectively). These results confirm that obesity is associated with decreased Na+, K+ -ATPase in intact red blood cells, and provide the first demonstration of a reduced sodium pump in isolated red cell membrane preparations from obese men.     

Alcohol decreases RhoA activity through a nitric oxide (NO)/cyclic GMP(cGMP)/protein kinase G (PKG)-dependent pathway in the airway epithelium

Background: Alcohol has been shown to have a number of harmful effects on the lung, including increasing the risk of pneumonia and bronchitis. How alcohol increases the risk of these diseases is poorly defined. RhoA is a small guanosine triphosphate (GTP)ase that plays an integral role in many basic functions of airway epithelial cells. It is not known how alcohol affects RhoA activity in the airway epithelium. We hypothesized that brief alcohol exposure modulates RhoA activity in the airway epithelium through a nitric oxide (NO)/cyclic GMP (cGMP)/protein kinase G (PKG)‐dependent pathway. Methods: Primary airway epithelial cells were cultured and exposed to ethanol at various concentrations and times. The cell layers were harvested and RhoA activity was measured. Results: Alcohol induced a time‐ and concentration‐dependent decrease in RhoA activity in airway epithelial cells. We were able to block this decrease in activity using Nω‐nitro‐l‐arginine methyl ester (L‐NAME) hydrochloride, a nitric oxide synthase (NOS) inhibitor. Likewise, we were able to demonstrate the same decrease in RhoA activation using 0.1 μM sodium nitroprusside, an NO donor. To determine the role of cGMP/PKG, we pretreated the cells with a cGMP antagonist analog, Rp‐8Br‐cGMPS. This blocked the decrease in RhoA activity caused by alcohol, suggesting that alcohol exerts its effect on RhoA activity through cGMP/PKG. Conclusions: Alcohol decreases airway epithelial RhoA activity through an NO/cGMP/PKG‐dependent pathway. RhoA activity controls many aspects of basic cellular function, including cell morphology, tight junction formation, and cell cycle progression and gene regulation. Dysregulation of RhoA activity can potentially have several consequences, including dysregulation of inflammation. This may partially explain how alcohol increases the risk of pneumonia and bronchitis.     

Studies of relationship among bile-acid uptake, Na+, K+-ATPase, and Na+ gradient in isolated cells from rat ileum

Studies were performed to determine relationships among Na+, K+-ATPase, the transmucosal Na+ gradient, and bile-acid transport in metabolically viable cells isolated from rat ileum. Incubation of cells with 0, 10(-6), 10(-5), 10(-4) and 10(-3) M ouabain resulted, respectively, in a 0, 10.3, 42.1, 97.0, and 100% decrease in glycocholate uptake and a 0, 10.7, 46.4, 76.8, and 100% decrease in Na+, K+-ATPase activity. Thus, one-half maximal inhibition of glycocholate uptake and Na+, K+-ATPase activity occurred at 5.5 x 10(-5) M and 1.7 x 10(-5) M ouabain, respectively. A change in glycocholate uptake was correlated with a change in Na+, K+-ATPase activity after daily injections of methylprednisolone. After 4 days treated animals showed a 26% and 36% increase in glycocholate uptake and Na+, K+-ATPase activity, respectively, over pair-fed saline-treated controls (p less than 0.001). Methylprednisolone did not significantly alter the activity of (Mg++)-ATPase when compared with controls (p greater than 0.05). Glycocholate uptake was reduced by the omission of Na+ from the incubation medium. Preincubation of cells at 37 degrees C with gramicidin D, 10 micrograms/ml, to alter membrane permeability to Na+, resulted in a significant rise in cell Na+ (p less than 0.01) and a significant fall in glycocholate uptake from values in untreated cells (p less than 0.01) to approach values for glycocholate uptake at 0 degrees C. These data suggest that Na+, K+-ATPase may play a role in a bile-acid uptake into ileal cells possibly by maintaining a Na+ electrochemical potential gradient for coupled Na+-bile-acid transport.     

Increases in Na+,K+-ATPase activity of erythrocytes and skeletal muscle after chronic ethanol consumption: evidence for reduced efficiency of the enzyme.

Increased Na+,K+-ATPase activity observed after chronic ethanol consumption has been examined to determine whether the increase is due to changes in the kinetic properties of the enzyme or increases in the amount of enzyme in the membranes examined. In skeletal muscle and erythrocyte ghosts from rat, as well as from humans, increased Na+,K+-ATPase activity in ethanol-consuming individuals was not accompanied by an increase in the number of ouabain binding sites. In studies with intact human erythrocytes, similar ouabain-sensitive 22Na+ and 86Rb+ pumping rates were observed between normal and ethanol-consuming individuals and the Na+ to Rb+ pumping ratio was found to be 1.5 in all cases. However, ouabain-sensitive lactate plus Pi formation was increased in cells from alcoholic individuals. Thus these data suggest that increased enzyme activity may be due to a kinetic alteration of the Na+,K+-ATPase and that the enzyme may be less efficient in coupling ion pumping to ATP hydrolysis than the enzyme in normal cells.     

Tissue specificity, localization in brain, and cell-free translation of mRNA encoding the A3 isoform of Na+,K+-ATPase.

The isolation of multiple Na+,K+-ATPase cDNAs from rat brain has led to the discovery of a family of alpha-isoform genes. Using A1 (alpha), A2 (alpha+), and A3 (alpha III) Na+,K+-ATPase gene probes, we have analyzed the distribution of Na+,K+-ATPase mRNAs in adult and fetal rat tissues by RNA blot and hybridization histochemistry. A1 Na+,K+-ATPase mRNA was found ubiquitously among various tissues, with highest levels in transport epithelial and neural tissues. A2 mRNA was found in adult neural and muscle tissues, and A3 mRNA was found only in neural tissues and fetal heart muscle. Both A1 and A2 mRNAs were less abundant in fetal brain than in adult brain; in contrast, A3 mRNA was abundant at both stages. In situ mapping of brain areas that contain A3 mRNA suggests that this Na+,K+-ATPase isoenzyme is expressed predominantly by neural cells. Analysis of Na+,K+-ATPase proteins generated by cell-free translation of synthetic mRNAs suggests that the A3 protein has properties similar to A2 (alpha+).     

Reduced Na++K+-ATPase activity in peripheral nerve of streptozotocin-diabetic rats: a role for protein kinase C?

Summary Six weeks after induction of diabetes, the rate of ouabain-sensitive ⁸⁶Rb⁺ accumulation, a parameter which reflects Na⁺+K⁺-ATPase pumping activity, was significantly reduced in endoneurial preparations of sciatic nerve from untreated diabetic rats compared with that in control rats (Trial 1, 0.19±0.09 versus 0.48±0.13 pmol/min per mg wet weight of tissue, p<0.001; Trial 2, 0.27±0.16 versus 0.47±0.18, p<0.01). This decrease in ouabain-sensitive ⁸⁶Rb⁺ uptake was not observed in nerves from diabetic rats maintained on sorbinil (an aldose reductase inhibitor) or myo-inositol diets. Protein kinase C activity was demonstrated in the soluble fraction of a sciatic nerve homogenate by assaying for lipid-activated, Ca⁺-dependent phosphorylation of calf thymus histone. No significant difference in the time course of kinase C activity was observed between cytosol fractions of nerve homogenates from control and diabetic rats (control, 6.22±0.97 pmol ³²P incorporated/mg cytosol protein in 50 min; diabetic, 5.32±0.71). Three low molecular weight neural proteins (each with M_r<29,000) were identified as substrates for protein kinase C.     

Development of an animal model for gestational hypermethioninemia in rat and its effect on brain Na+,K+-ATPase/Mg2+-ATPase activity and oxidative status of the offspring

In the present study we developed a chemically induced experimental model for gestational hypermethioninemia in rats and evaluated in the offspring the activities of Na⁺,K⁺-ATPase and Mg²⁺-ATPase, as well as oxidative stress parameters, namely sulfhydryl content, thiobarbituric acid-reactive substances and the antioxidant enzymes superoxide dismutase and catalase in encephalon. Serum and encephalon levels of methionine and total homocysteine were also evaluated in mother rats and in the offspring. Pregnant Wistar rats received two daily subcutaneous injections of methionine throughout the gestational period (21 days). During the treatment, a group of pregnant rats received dose 1 (1.34 μmol methionine/g body weight) and the other one received dose 2 (2.68 μmol methionine/g body weight). Control group received saline. After the rats give birth, a first group of pups was killed at the 7th day of life and the second group at the 21th day of life for removal of serum and encephalon. Mother rats were killed at the 21th day postpartum for removal of serum and encephalon. Both doses 1 and 2 increased methionine levels in encephalon of the mother rats and dose 2 increased methionine levels in encephalon of the offspring. Maternal hypermethioninemia also decreased the activities of Na⁺,K⁺-ATPase, Mg²⁺-ATPase and catalase, as well as reduced total sulfhydryl content in the encephalon of the pups. This chemical model seems to be appropriate for studies aiming to investigate the effect of maternal hypermethioninemia on the developing brain during gestation in order to clarify possible neurochemical changes in the offspring.     

Platelet Na+,K+-ATPase activity as a possible peripheral marker for the neurotoxic effects of phenylalanine in phenylketonuria

Thein vitro effects of phenylalanine or alanine alone or combined on Na⁺,K⁺-ATPase activity in membranes from human platelets were investigated. The enzyme activity was assayed in membranes prepared from platelet-rich plasma of healthy donors. Phenylalanine or alanine were added to the assay to final concentrations of 0.3 to 1.2 mM, similar to those found in plasma of phenylketonuric patients. Phenylalanine inhibited Na⁺,K⁺-ATPase activity by 20–50% [F(4,25)=11.47; p<0.001]. Alanine had no effect on Na⁺,K⁺-ATPase activity but when combined with phenylalanine prevented the enzyme inhibition. These results, allied to others previously reported on brain Na⁺,K⁺-ATPase activity, may reflect a general inhibitory effect of phenylalanine on this important enzyme activity. Therefore, it is possible that measurement of Na⁺,K⁺-ATPase activity in platelets from PKU patients may be a useful peripheral marker for the neurotoxic effects of phenylalanine.     

Effect of phenylalanine and p-chlorophenylalanine on Na+, K+-ATPase activity in the synaptic plasma membrane from the cerebral cortex of rats

Na⁺, K⁺-ATPase activity was measured in synaptic plasma membrane from cerebral cortex of Wistar rats subjected to experimental phenylketonuria, i.e., chemical hyperphenylalaninemia induced by subcutaneous administration of 5.2 μmol phenylalanine /g body weight (twice a day) plus 0.9 μmol p-chlorophenylalanine /g body weight (once a day). The treatment was performed from the 6^th to the 14^th postpartum day and rats were killed 12 h after the last injection. Synaptic plasma membrane from cerebral cortex was prepared by a discontinuous density sucrose gradient for Na⁺, K⁺-ATPase activity determination. The results showed that the enzyme activity was decreased by 30% in animals subjected to experimental phenylketonuria when compared to control. Thein vitro effects of the drugs on Na⁺, K⁺-ATPase activity were also investigated. Phenylalanine and p-chlorophenylalanine inhibited the enzyme activity and this inhibition was reversed by alanine. In addition, competition between phenylalanine and p-chlorophenylalanine for binding to the enzyme was observed, suggesting a common binding site for these substances. Our results suggest that reduction of Na⁺, K⁺-ATPase activity may be one of the mechanisms related to the brain dysfunction observed in human PKU.     

冠心病患者血小板PKC、PKCI及红细胞PKC活性变化

目的 研究蛋白激酶C(PKC)在冠心病的发生发展中的作用。方法 测定87例心绞痛(AP)患者、91例急性心肌梗死(AMl)患者、90例健康对照(HC)者的血小板胞膜、胞浆PKC、胞浆蛋白激酶C抑制剂(PKCl)活性和红细胞胞膜、胞浆PKC活性。结果 AP组和AMI组血小板胞膜中PKC活性明显高于HC组,而胞浆中PKC活性明显低于HC组;AP组和AMI组血小板胞浆中PKCI活性明显低于HC组;AP组和AMI组红细胞胞膜中PKC活性明显高于HC组,而胞浆中PKC活性低于HC组。结论 PKC可能参与冠心病的发病。     

The decreased phosphorylation of cyclic adenosine monophosphate (cAMP) response element binding (CREB) protein in the central amygdala acts as a molecular substrate for anxiety related to ethanol withdrawal in rats

BACKGROUND: Several lines of evidence indicate a high comorbidity between anxiety and alcohol abuse. This study investigated the molecular mechanisms in the amygdaloid neurocircuitry governing anxiety related to ethanol withdrawal and also the phenomenon of alcohol preference. METHODS: Male Sprague Dawley(R) rats were treated with ethanol or control diet for 15 days, and ethanol-fed rats were withdrawn for 0 and 24 hr. Ethanol-withdrawn or control diet-fed rats were bilaterally infused into central or basolateral amygdala with artificial cerebrospinal fluid or protein kinase A (PKA) activator or inhibitor. These rats were used to measure anxiety levels by the elevated plus-maze test. Protein levels of various signaling molecules related to cyclic adenosine monophosphate (cAMP)-response element binding (CREB) protein signaling in amygdaloid structures were determined by gold immunolabeling procedure. The messenger RNA levels of neuropeptide Y were determined by in situ polymerase chain reaction procedure. RESULTS: Ethanol withdrawal (24 hr) after chronic exposure (15 days) produced anxiety in rats as measured by elevated plus-maze test. Ethanol withdrawal but not treatment significantly decreased the phosphorylation of CREB protein and protein levels of Ca2+/calmodulin-dependent protein kinase IV without modulating the protein levels of total CREB and alpha-catalytic subunit of protein kinase A (PKA-Calpha) in the central and medial amygdala. However, these changes were not observed in the basolateral amygdala. We also investigated the effects of manipulation of the phosphorylation status of CREB in the central amygdala by infusion of the PKA activator (Sp-cAMPS) or inhibitor (Rp-cAMPS) on anxiety levels in rats during ethanol withdrawal. When Sp-cAMPS is specifically infused into the central amygdala, it dose-dependently normalizes the decrease in CREB phosphorylation and prevents the development of anxiety in rats during ethanol withdrawal. On the other hand, Rp-cAMPS infusions into the central or basolateral amygdala decrease CREB phosphorylation, but only infusion into the central amygdala provokes anxiety and increases alcohol preference in normal rats. We also found that alcohol preference provoked by decreased CREB phosphorylation is related to decreased expression of the neuropeptide Y gene in the central amygdala. CONCLUSIONS: These novel results suggest the possibility that decreased CREB phosphorylation in the central amygdala acts as a common molecular correlate for anxiety and alcohol-drinking behaviors and also is correlated with anxiety related to ethanol withdrawal.     

Decrease in Na+,K+-ATPase activity and [3H]ouabain binding sites in sarcolemma prepared from hearts of spontaneously hypertensive rats

Na+,K+-ATPase activity, phosphorylation, and [3H]ouabain binding in sarcolemma isolated from spontaneously hypertensive rat (SHR) hearts were compared to the same parameters in sarcolemma from normotensive rat (WKY) hearts. Sarcolemma prepared from SHR heart contained significantly less ouabain-inhibitable ATPase activity than sarcolemma from WKY heart. No significant differences in sarcolemmal protein content or recovery were noted between the two groups. The numbers of phosphorylation sites and ouabain binding sites were lower for SHR hearts than for WKY hearts. The KD values for ouabain binding were the same (0.30 muM) in cardiac sarcolemma of SHR and WKY. The I50 values for inhibition by ouabain of Na+,K+-ATPase were also the same for both groups (SHR = 49 microM; WKY = 44 microM). These data suggest that the decrease of cardiac sarcolemmal Na+,K+-ATPase activity in SHR hearts is due to a decrease in the number of active sites.     

Sphingosine and psychosine, suggested inhibitors of protein kinase C, inhibit LH effects in rat luteal cells

The possible involvement of protein kinase C on luteinizing hormone (LH) effects in dispersed rat luteal cells was investigated using two substances that have been reported to be protein kinase C inhibitors, sphingosine and psychosine. Sphingosine efficiently inhibited protein kinase C activity both in brain and luteal cytosol fractions. Both substances inhibited LH-stimulated cyclic adenosine monophosphate (cAMP) accumulation in a dose-dependent fashion with an ID₅₀ at 3–7 μM (sphingosine) and 40 μ.m (psychosine). LH-stimulated progesterone production was also inhibited with an ID₅₀ at 6–10 μM (sphingosine) and 40–100 μM (psychosine). The inhibition was not due to an increased phosphodiesterase activity since IBMX (3-isobutyl-1-methylxanthine, 0.1 mM) and RO 20–1724 (4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone, 0.1 mM) did not abolish the inhibitory effect of sphingosine. To study the mode of action of sphingosine, forskolin and cAMP analogues were tested. The effect of these substances on Steroidogenesis was inhibited, as well as the forskolin-induced cAMP accumulation, by sphingosine.

This study demonstrates a clear inhibition of LH-stimulated effects by sphingosine and psychosine. LH action in rat luteal cells is discussed in relation to protein kinase C and the possible mode of sphingosine action.