Effects in vitro of copper and zinc on hepatic cytochrome P-450 activities

1. In previous studies we have shown that hepatic copper and zinc increases and liver microsomal cytochrome P-450 activities greatly decreases in adjuvant arthritic rats. 2. In the present paper we study if the changes in copper and zinc could be related to depression of drug microsomal activity. Thus, the effect of in vitro addition of copper or zinc to microsomal fraction upon aminopyrine N-demethylase (AND) and aniline p-hydroxylase (APH) activity was measured. 3. Both metals produced an inhibition of enzyme activity. The reduction of AND and APH activities produced by copper (ID25 = 4.7 x 10(-5)M to AND; 1.05 x 10(-5)M to APH) was greater than that obtained with zinc (ID25 = 2.26 x 10(-4)M to AND; 3.3 x 10(-4)M to APH).     

The effect of metoclopramide on inhibition induced by purine nucleotides, noradrenaline, and theophylline ethylenediamine on intestinal muscle and on peristalsis in vitro.

Metoclopramide (N-(diethylaminoethyl)-2-methoxy-4-amino-5-chlorobenzamide) (Mcp) at concentrations of 0.1 and 1.0 muM partially and significantly reduced the relaxations induced by adenosine 5'-triphosphate (ATP), adenosine diphosphate (ADP), and adenosine, was without effect on theophylline ethylenediamine whilst significantly potentiating noradrenaline on the atropine-pretreated (0.1 muM) taenia coli, rabbit ileum, and rat duodenum. Mcp (1.0 muM) decreased the inhibitory effects of ATP, ADP, and adenosine on peristalsis induced in the isolated guinea-pig ileum by a constant increase in intraluminal pressure, did not affect inhibition due to theophylline ethylenediamine, whilst it potentiated inhibition of peristalsis due to noradrenaline. It is proposed that this effect of Mcp may be a specific antagonistic action on receptors sensitive to the putative purinergic transmitter, ATP and ADP, and may be partly responsible for its observed facilitatatory action on peristalsis.     

Pleurotus ostreatus, an oyster mushroom, decreases the oxidative stress induced by carbon tetrachloride in rat kidneys, heart and brain

The present work is aimed at evaluating the protective effect of the oyster mushroom, Pleurotus ostreatus on carbon tetrachloride (CCl4)-induced toxicity in male Wistar rats. Significantly elevated mean levels (p<0.05) of malondialdehyde (MDA) and lowered mean levels (p<0.01) of reduced glutathione (GSH), vitamins C and E (p<0.05) were observed in kidneys, heart and brain of rats exposed to CCl4, when compared to values in normal rats. Quantitative and qualitative analysis of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (Gpx) and glutathione-S-transferase (GST) revealed lower activities of these antioxidant enzymes in the kidneys, heart and brain of rats exposed to CCl4. When the extract of P. ostreatus was used to treat rats with CCl4-induced toxicity, it lowered the mean level of MDA, elevated the mean levels of GSH and of vitamins C and E and enhanced the mean activities of CAT, SOD, Gpx and GST so that the values of most of these parameters did not differ significantly from those of normal rats. Histopathological studies confirmed the toxic effects of CCl4 on other organs such as kidneys, heart and brain and also tissue protective effect of the extract of P. ostreatus. These results suggest that an extract of P. ostreatus is able to alleviate the oxidative damage caused by CCl4 in the kidneys, heart and brain of Wistar rats.     

Moderate hypoxia: reactivity of pial arteries and local effect of theophylline

The reactivity of pial arteries to the perivascular microapplication of artificial cerebrospinal fluids with mounting concentrations of adenosine (10(-11)-10(-3) M), K+ (0-10 mM), and H+ (pH 5.1-7.6) was determined in chloralose-anesthetized ventilated cats during normoxic control conditions and during moderate normocapnic arterial hypoxia (arterial Po2 47 Torr). Hypoxia induced a significant mean pial arterial dilatation of 18-29% in the various types of experiments. The pial arterial reactivity to each of the tested factors remained unchanged during hypoxia compared with normoxia. The hypoxic vasodilatation could not be reduced by the perivascular microapplication of theophylline (10(-5) and 5 X 10(-5) M). Systemic theophylline (50-75 mumol/kg, iv), regardless of whether given during or before hypoxia, did not attenuate the hypoxic vasodilatation, although it blocked dilatations induced by the perivascular microapplication of adenosine during normoxia. The present study shows that 1) local metabolic factors are vasoactive during moderate hypoxia; therefore they could mediate the hypoxic dilatation of brain vessels; 2) systemic theophylline can block vascular adenosine receptors; 3) since local theophylline had no effect on the hypoxic dilatation of pial arteries, adenosine may not be the main causative factor for the hypoxic hyperemia.     

Theophylline stimulation of gluconeogenesis from alanine in normal and sympathectomized rats.

The influence of theophylline ethylenediamine (100 mg/kg i.p.) on gluconeogenesis was studied in normal and in adrenodemedullated and reserpinized rats after overnight fasting by measuring the time-course of Alanine-14C incorporation into Glucose-14C. In the normal rat, theophylline produced a moderate hyperglycemia associated with an increased conversion of alanine to glucose at all time intervals. In addition, a marked rise of plasma levels of insulin and glucagon was observed. In sympathetctomized rats, plasma glucose and gluconeogenesis were again enhanced by theophylline, but the pattern of these modifications differed from that of normal rats since the peak values occurred earlier. Subsequently, both parameters rapidly declined reaching values lower than controls at the end of the experiment. Insulin response to theophylline was higher in sympathectomized animals in comparison to normal rats, while glucagon response was approximately of the same magnitude in the two groups. From these findings it was concluded that theophylline is able to stimulate gluconeogenesis from alanine both in the normal and sympathectomized rat. The different pattern of alanine conversion to glucose seems to depend on the different participation of insulin and catecholamines in the two groups.     

Regulation of cyclic AMP level and lactic acid production in Ehrlich ascites tumor cells.

1. Quercetin (3.3',4',5,7-pentahydroxy flavone) at the concentration of 10(-4) M, as well as 2-10(-2) M theophylline and 1.5 - 10(-4) M prostaglandin E2 caused maximal rise of cyclic AMP in Ehrlich ascites tumor cells. 2. No additional increase of cyclic AMP level in these cells was found when both quercetin (10(-4) M) and theophylline (2-10(-2) M) were present in the incubation medium, while combination of quercetin (10(-4) M) and prostaglandin E2 (1.5 - 10(-4) M) has a synergistic effect on the level of cyclic AMP. 3. Degradation of cyclic AMP by homogenate of Ehrlich ascites tumor cells was inhibited by both quercetin and theophylline. 4. Quercetin, and to a smaller but significant extent theophylline, inhibited the lactic acid production in Ehrlich ascites tumor cells while prostaglandin E2 did not change the glycolytic rate in these cells. No synergistic inhibitory effect on lactic acid production was found when combinations of quercetin and prostaglandin E2, quercetin and theophylline or prostaglandin E2 and theophylline were tested. 5. Treatment of Ehrlich ascites tumor cells with dextran sulfate abolished the inhibitory effect of quercetin on lactic acid production, while the effect of the bioflavonoid on cyclic AMP levels was not altered.     

Protection of calcitonin gene-related peptide-mediated preconditioning against coronary endothelial dysfunction induced by reperfusion in the isolated rat heart

This study was designed to explore the protective effect of ischemic preconditioning on reperfusion-induced coronary endothelial dysfunction, with a focus on the role of calcitonin gene-related peptide (CGRP) in this effect, in the isolated perfused rat heart. Thirty minutes of global ischemia and 30 min of reperfusion significantly decreased heart rate, left ventricular pressure, and its first derivative and impaired vasodilator responses to acetylcholine. Ischemia-reperfusion did not affect vasodilator responses to sodium nitroprusside. Preconditioning induced by three cycles of 5 min of ischemia and 5 min of reperfusion produced a significant improvement in cardiac function concomitantly with an amelioration of vasodilator responses to acetylcholine. The protective effects of ischemic preconditioning were abolished by CGRP(8-37) (10(-7) M) , the selective CGRP receptor antagonist. After pretreatment with capsaicin (50 mg x kg(-1), s.c.) to deplete endogenous CGRP, the preconditioning effect was absent. Pretreatment with exogenous CGRP (5 x 10(-9) M) for 5 min induced a preconditioning-like protection. The present study suggests that the cardioprotection of ischemic preconditioning is related to the preservation of the coronary endothelial cell, and that the protective effect of preconditioning is mediated by endogenous CGRP in the isolated perfused rat heart.     

Are goose nesting success and lemming cycles linked? Interplay between nest density and predators

The suggested link between lemming cycles and reproductive success of arctic birds is caused by potential effects of varying predation pressure (the Alternative Prey Hypothesis, APH) and protective association with birds of prey (the Nesting Association Hypothesis, NAH). We used data collected over two complete lemming cycles to investigate how fluctuations in lemming density were associated with nesting success of greater snow geese ( Anser caerulescens atlanticus ) in the Canadian High Arctic. We tested predictions of the APH and NAH for geese breeding at low and high densities. Goose nesting success varied from 22% to 91% between years and the main egg predator was the arctic fox ( Alopex lagopus ). Nesting associations with snowy owls ( Nyctea scandiaca ) were observed but only during peak lemming years for geese nesting at low density. Goose nesting success declined as distance from owls increased and reached a plateau at 550 m. Artificial nest experiments indicated that owls can exclude predators from the vicinity of their nests and thus reduce goose egg predation rate. Annual nest failure rate was negatively associated with rodent abundance and was generally highest in low lemming years. This relationship was present even after excluding goose nests under the protective influence of owls. However, nest failure was inversely density-dependent at high breeding density. Thus, annual variations in nest density influenced the synchrony between lemming cycles and oscillations in nesting success. Our results suggest that APH is the main mechanism linking lemming cycles and goose nesting success and that nesting associations during peak lemming years (NAH) can enhance this positive link at the local level. The study also shows that breeding strategies used by birds (the alternative prey) could affect the synchrony between oscillations in avian reproductive success and rodent cycles.     

Kinetic mechanism of enterococcal aminoglycoside phosphotransferase 2"-Ib

The major mechanism of resistance to aminoglycosides in clinical bacterial isolates is the covalent modification of these antibiotics by enzymes produced by the bacteria. Aminoglycoside 2'-Ib phosphotransferase [APH(2')-Ib] produces resistance to several clinically important aminoglycosides in both Gram-positive and Gram-negative bacteria. Nuclear magnetic resonance analysis of the product of kanamycin A phosphorylation revealed that modification occurs at the 2'-hydroxyl of the aminoglycoside. APH(2')-Ib phosphorylates 4,6-disubstituted aminoglycosides with kcat/Km values of 10(5)-10(7) M-1 s-1, while 4,5-disubstituted antibiotics are not substrates for the enzyme. Initial velocity studies demonstrate that APH(2')-Ib operates by a sequential mechanism. Product and dead-end inhibition patterns indicate that binding of aminoglycoside antibiotic and ATP occurs in a random manner. These data, together with the results of solvent isotope and viscosity effect studies, demonstrate that APH(2')-Ib follows the random Bi-Bi kinetic mechanism and substrate binding and/or product release could limit the rate of reaction.     

Free radical scavenging is involved in the protective effect of L-propionyl-carnitine against ischemia-reperfusion injury of the heart.

L-Propionyl-carnitine is known to improve the recovery of myocardial function and metabolic parameters reduced in the course of ischemia-reperfusion of the heart. The mechanism of this protective effect of L-propionyl-carnitine is not fully understood. The purpose of this study was to elucidate the effects of L-propionyl-carnitine in Langendorff perfused rat hearts subjected to 40 min of ischemia followed by 20 min of reperfusion. We tested the hypothesis that L-propionyl-carnitine suppresses generation of oxygen radicals and subsequent oxidative modification of myocardial proteins during reperfusion. Our data show that the protective effect of L-propionyl-carnitine in the course of ischemia-reperfusion is highly significant in terms both of mechanical properties of the heart (developed pressure) and of high-energy phosphates (ATP, creatine phosphate). Myocardial creatine phosphokinase (CPK) activity decreased in the course of the reperfusion period. The loss of CPK activity was partially prevented by L-propionyl-carnitine. Two other effects were observed when L-propionyl-carnitine was present in the perfusion solution: (i) the reperfusion-induced sharp increase in oxidative protein modification was completely prevented as detected by the formation of protein carbonyls, and (ii) generation of hydroxyl radicals was significantly inhibited as detected by the formation of the adducts with the spin trap 5,5-dimethyl-1-pyrroline-1-oxide. We conclude that the protective effect of L-propionyl-carnitine against ischemia-reperfusion injury of the heart is at least due in part to its ability to suppress the development of oxidative stress and free radical damage.     

Cyclic nucleotides and calcium: their role in the control of cell communication in the heart

The effects of theophylline and di-butyryl cyclic adenosine 3',5'-monophosphate (db-cAMP) on the electrical coupling of heart cells were investigated in rat trabeculae. Theophylline (4 X 10(-4) M) and db-cAMP (5 X 10(-5) M) increased both the space constant and conduction velocity. The time constant of the membrane was not changed by either drug. Measurements of the time constant of the foot of the action potential and conduction velocity were used to calculate the intracellular longitudinal resistance. Both theophylline and db-cAMP were found to enhance cell-to-cell communication in the heart by decreasing the intracellular longitudinal resistance.     

Structure and function of APH(4)-Ia, a hygromycin B resistance enzyme

The aminoglycoside phosphotransferase (APH) APH(4)-Ia is one of two enzymes responsible for bacterial resistance to the atypical aminoglycoside antibiotic hygromycin B (hygB). The crystal structure of APH(4)-Ia enzyme was solved in complex with hygB at 1.95 Å resolution. The APH(4)-Ia structure adapts a general two-lobe architecture shared by other APH enzymes and eukaryotic kinases, with the active site located at the interdomain cavity. The enzyme forms an extended hydrogen bond network with hygB primarily through polar and acidic side chain groups. Individual alanine substitutions of seven residues involved in hygB binding did not have significant effect on APH(4)-Ia enzymatic activity, indicating that the binding affinity is spread across a distributed network. hygB appeared as the only substrate recognized by APH(4)-Ia among the panel of 14 aminoglycoside compounds. Analysis of the active site architecture and the interaction with the hygB molecule demonstrated several unique features supporting such restricted substrate specificity. Primarily the APH(4)-Ia substrate-binding site contains a cluster of hydrophobic residues that provides a complementary surface to the twisted structure of the substrate. Similar to APH(2″) enzymes, the APH(4)-Ia is able to utilize either ATP or GTP for phosphoryl transfer. The defined structural features of APH(4)-Ia interactions with hygB and the promiscuity in regard to ATP or GTP binding could be exploited for the design of novel aminoglycoside antibiotics or inhibitors of this enzyme.     

Protective effects of N-acetyl cysteine on lipid peroxide metabolism on isoproterenol-induced myocardial infarcted rats

The present study was designed to evaluate the preventive effects of N-acetyl cysteine on lipid peroxide metabolism in isoproterenol (ISO) induced myocardial infarcted rats. Male albino Wistar rats were pretreated with N-acetyl cysteine (5 and 10 mg/kg) daily for a period of 14 days. After the pretreatment period, ISO (100 mg/kg) was subcutaneously injected to rats twice at an interval of 24 h. Increased activities of serum creatine kinase, creatine kinase-MB, lactate dehydrogenase, and increased intensities of serum lactate dehydrogenase-isoenzyme bands (LDH-1, LDH-2) were observed in ISO-induced rats. The heart lipid peroxidation products were significantly increased, and the antioxidant system was significantly reduced in ISO-induced rats. Pretreatment with N-acetyl cysteine (5 and 10 mg/kg) to ISO-induced rats showed significant effects on all the biochemical parameters studied. Histopathological findings of the myocardium also showed the protective role of N-acetyl cysteine in ISO-induced rats. Furthermore, in vitro study confirmed the potent-free radical scavenging activity of N-acetyl cysteine. The effect at a dose of 10 mg/kg of N-acetyl cysteine was more pronounced than the dose, 5 mg/kg. The results of our study show that N-acetyl cysteine protects the heart against ISO-induced myocardial infarction by its free radical scavenging effect.     

Inosine Mediates the Protective Effect of Adenosine in Rat Astrocyte Cultures Subjected to Combined Glucose-Oxygen Deprivation

Abstract: Preliminary evidence suggests adenosine, a neuromodulator, has neuroprotective properties during cerebral ischemia. It is unclear, however, if adenosine has glioprotective effects. We studied the effect of adenosine on cellular injury in astroglial cultures subjected to combined glucose-oxygen deprivation. Adenosine (100–1,000 µ M ) dramatically reduced astroglial injury, whereas the adenosine agonists 2-chloroadenosine (10 n M –100 µ M ), N ⁶-cyclopentyladenosine (1 n M –10 µ M ), 5'- N -ethylcarboxamidoadenosine (10 n M –100 µ M ), and N ⁶-2-(4-aminophenyl)ethyladenosine (10 n M –100 µ M ) had no effect. Furthermore, the adenosine antagonists 8-cyclopentyl-1,3-dipropylxanthine (1 n M –1 µ M ), xanthine amine congener (10 n M –10 µ M ), and 8-( p -sulfophenyl)-theophylline (10–300 µ M ) failed to reverse the protective effect of 200 µ M adenosine. Next, adenosine degradation products were studied. Inosine proved to be glioprotective at concentrations nearly identical to those of adenosine, but hypoxanthine and ribose had no effect. The protective effect of 200 µ M inosine was not reversed by 8-( p -sulfophenyl)theophylline (10–300 µ M ). Adenosine deaminase (1 unit/ml) had no effect on protection produced by adenosine, whereas erythro -9-(2-hydroxy-3-nonyl)adenine hydrochloride (10 µ M ) reversed the protective effect of adenosine. Dipyridamole (4 µ M ) inhibited the protective effect of both adenosine and inosine. We conclude that adenosine dramatically decreases astroglial injury during combined glucose-oxygen deprivation and that this protective effect appears to be mediated by inosine.     

Treatment of nocturnal asthma: the role of sustained-release theophylline and oral beta-2-mimetics

In two double-blind, multiple-dose cross-over studies the therapeutic effects of SR theophylline preparations given once each night (mean 11.2 mg/kg per day) versus twice daily in equal doses (mean 10.3 mg/kg per day) (study I) and SR-terbutaline in equal doses (mean 0.25 mg/kg per day) versus SR theophylline in unequally divided daily doses (mean 5.3 mg/kg morning dose, 10.6 mg/kg evening dose) study II) were compared in 19 patients with nocturnal asthma. At the end of each treatment period drug serum concentrations and PEFR were measured every 2 hr over a 24-hr period. With the twice-daily, equally divided regimen, serum theophylline concentrations were lower at night than during the day (mean 9.4 +/- 0.9 versus 11.3 +/- 1.0 mg/l). With the single evening administration, serum theophylline concentrations were considerably higher at night (Cmax 16.3 +/- 1.4 mg/l) and the circadian variation of PEFR was significantly reduced. PEFR was higher during night and early morning (283 +/- 14 versus 217 +/- 11 l/min, P less than 0.005). During daytime in study II, PEFR values were slightly higher with theophylline than terbutaline. There was no significant difference in peak flow between either treatment during the night and early morning. However, additional use of inhaled beta-2-mimetics because of asthmatic attacks occurred more often during terbutaline (79 times in 8/10 patients) than theophylline treatment (29 times in 5/10 patients). Symptom scores, number of attacks and side-effects clearly favor the theophylline regimen. We conclude that for patients with nocturnal asthma a once-nightly dose of SR theophylline can be sufficient for stabilization of the airways.     

Increasing intracellular cAMP and cGMP inhibits cadmium-induced oxidative stress in rat submandibular saliva

The effect of cadmium on induction of oxidative stress in rat submandibular saliva and protective role of increasing intracellular cAMP and cGMP by use of specific phosphodiesterase inhibitors, theophylline and sildenafil were investigated. Pure submandibular saliva was collected intraorally by micro polyethylene cannula from anaesthetized rats using pilocarpine as secretagogue. Acute administration of cadmium (10 mg/kg) caused significant oxidative stress by increasing lipid peroxidation by-products (thiobarbituric reactive substances, TBARS) and decreasing total thiols and total antioxidant power of the saliva. Concurrent therapy of rats by theophylline (25 mg/kg) and sildenafil (5 mg/kg) prevented cadmium-induced oxidative stress in saliva. Theophylline and sildenafil inhibited cadmium-induced increase in lipid peroxidation and decrease in total thiols and antioxidant power. It is concluded that cadmium administration results in oxidative stress in rat submandibular saliva, which can be protected by concurrent administration of specific cyclic nucleotide phosphodiesterase inhibitors.     

Concentration-dependent interaction of theophylline with d-tubocurarine

The interaction of theophylline with d-tubocurarine chloride (dTC) was examined in rabbits. After steady-state subtherapeutic (less than 10 mg/l), therapeutic (10-20 mg/l), and toxic (greater than 20 mg/l) concentrations of theophylline, dose-response curves for dTC were determined and compared with controls that received no theophylline. At therapeutic concentrations of theophylline the effective dose for 50% inhibition of twitch (ED50) for dTC (mean +/- SE, 0.115 +/- 0.016 mg/kg) was significantly shifted to the left in comparison with the control (0.165 +/-0.008 mg/kg). The ED50 of dTC for the subtherapeutic group was 0.143 +/- 0.011 mg/kg, which was less than the control but not of statistical significance (P = 0.1). The ED50 for the toxic theophylline group was 0.168 +/- 0.003 mg/kg, which was not significantly different from controls but significantly different from the theophylline therapeutic and subtherapeutic groups. Thus, toxic concentrations of theophylline reversed the potentiating effects of therapeutic and subtherapeutic concentrations of dTC dose-response curves. Therefore, depending on concentration, theophylline exhibits a biphasic interaction with dTC. Surgical patients on theophylline may require less dTC intraoperatively. More importantly, the use of theophylline in the postoperative period to reverse anesthetic effects may result in recurarization.     

Oxygen Radical Injury and Loss of High-Energy Compounds in Anoxic and Reperfused Rat Heart: Prevention By Exogenous Fructose-1, 6-Bisphosphate

Isolated Langendorff-perfused rat hearts after 10 minutes preperfusion, were subjected to a substrate-free anoxic perfusion (20 minutes) followed by 20 minutes reperfusion with a glucose-containing oxygen-balanced medium. Under the same perfusion conditions, the effect of exogenous 5mM fructose-1, 6-bisphosphate has been investigated. The xanthine dehydrogenase to xanthine oxidase ratio, concentrations of high-energy phosphates and the TBA-reactive material (TBARS) were determined at the end of each perfusion period in both control and fructose-1, 6-bisphosphate-treated hearts. Results indicate that anoxia induces the irreversible transformation of xanthine dehydrogenase into oxidase as a consequence of the sharp decrease of the myocardial energy metabolism. This finding is supported by the protective effect exerted by exogenous fructose-1, 6-bisphosphate which is able to maintain the correct xanthine dehydrogenase/oxidase ratio by preventing the depletion of phosphorylated compounds during anoxia. Moreover, in control hearts, the release oflactate dehydrogenase during reperfusion, is paralleled by a 50% increase in the concentration of tissue TBARS. On the contrary, in fructose-1, 6-bisphosphate-treated hearts this concentration does not significantly change after reoxygenation, while a slight but significant increase of lactate dehydrogenase activity in the perfusates is observed.

On the whole these data indicate a direct contribution of oxygen-derived free radicals to the worsening of post-anoxic hearts. A hypothesis on the mechanism of action of fructose-1, 6-bisphosphate in anoxic and reperfused rat heart and its possible application in the clinical therapy of myocardial infarction are presented.     

Effect of cyclic AMP on cellular contractility and DNA synthesis in chorioretinal fibroblasts maintained in collagen matrices

Dibutyryl cyclic AMP (db-cAMP), theophylline and forskolin were found to be potent inhibitors of DNA synthesis and cell contractility in chorioretinal fibroblasts maintained in 3-dimensional collagen matrices. Dose-response curves were constructed for the inhibitory action of these agents on both cellular parameters and their interrelationship examined by regression analysis. The results obtained indicate that these parameters were equally inhibited by cAMP with the exception of high concentrations of db-cAMP (greater than 10(-3) M) where a greater effect on DNA synthesis than cell contractility was observed which was attributed to additional inhibition of DNA synthesis by db-cAMP degradation products. It is proposed from the present results and those of other investigators that cAMP regulates non-transformed fibroblast proliferation and contraction through a common regulatory mechanism possibly involving cAMP-dependent protein kinases and calcium ions.     

Effects of theophylline on expression of the long cilia phenotype in sand dollar blastulae

Previously, increases in ciliary length have only been obtained through genetic mutation in Chlamydomonas or by incubation of swimming echinoderm blastulae in trypsin or elastase. We have found that the phenotypic switch from short to long cilia on sand dollar blastulae can also be effected by incubation in theophylline. Cilia detached from control blastulae have a mean length of 21 +/- 7 microns with 10% of the cilia being greater than 30 microns. Upon incubation in 10 mM theophylline additional long cilia appeared after 10 hours and by 24-32 hours 1/2-3/4 of the embryo was covered with long cilia. The percentage of long cilia increased to 65% with a mean length of 40.0 +/- 17.6 microns. Incubation in other methylxanthines, such as aminophylline, caffeine, or isobutylmethylxanthine, inhibited development but had no effect on ciliary length distribution. Dibutyryl cAMP, 8-bromoadenosine, and calcium ionophore also had no effect on ciliary length. Cyclic AMP levels were measured and showed only slight differences among controls and embryos incubated in trypsin, caffeine, or theophylline. These data suggest that theophylline may be altering ciliary length control through some mechanism other than elevations in cAMP.