Demeclocycline attenuates hyponatremia by reducing aquaporin-2 expression in the renal inner medulla.

Binding of vasopressin to its type 2 receptor in renal collecting ducts induces cAMP signaling, transcription and translocation of aquaporin (AQP)2 water channels to the plasma membrane, and water reabsorption from the prourine. Demeclocycline is currently used to treat hyponatremia in patients with the syndrome of inappropriate antidiuretic hormone secretion (SIADH). Demeclocycline's mechanism of action, which is poorly understood, is studied here. In mouse cortical collecting duct (mpkCCD) cells, which exhibit deamino-8-D-arginine vasopressin (dDAVP)-dependent expression of endogenous AQP2, demeclocycline decreased AQP2 abundance and gene transcription but not its protein stability. Demeclocycline did not affect vasopressin type 2 receptor localization but decreased dDAVP-induced cAMP generation and the abundance of adenylate cyclase 3 and 5/6. The addition of exogenous cAMP partially corrected the demeclocycline effect. As in patients, demeclocycline increased urine volume, decreased urine osmolality, and reverted hyponatremia in an SIADH rat model. AQP2 and adenylate cyclase 5/6 abundances were reduced in the inner medulla but increased in the cortex and outer medulla, in the absence of any sign of toxicity. In conclusion, our in vitro and in vivo data indicate that demeclocycline mainly attenuates hyponatremia in SIADH by reducing adenylate cyclase 5/6 expression and, consequently, cAMP generation, AQP2 gene transcription, and AQP2 abundance in the renal inner medulla, coinciding with a reduced vasopressin escape response in other collecting duct segments.

Drug-induced teratogenesis in vitro: inhibition of calcification by different tetracyclines.

Inhibition of calcification in embryonic bone rudiments was studied in the presence of several tetracyclines at three different concentrations. Different criteria for calcification and different concentrations of tetracyclines yielded parallel results and showed significant differences in the inhibitory action of the various compounds. The clear-cut results indicate that the test-system that was developed may be useful for the comparison of various teratogens under simplified controllable conditions.

An in vitro study of the antimicrobial activity of some endodontic medicaments and their bases using an agar well diffusion assay.

BACKGROUND: The aim of this study was to determine the in vitro antimicrobial activities of various endodontic medicaments and their bases against selected organisms using an agar diffusion assay. METHODS: An agar well diffusion assay was used to test the antimicrobial action of some commonly used endodontic medicaments (Ledermix paste, Pulpdent paste, Ultracal paste, and a 50:50 mix of Ledermix and Pulpdent pastes) and their bases. Three bacterial species (E. faecalis, P. micros, P. intermedia) and one yeast (C. albicans) were selected. The diameters of growth inhibition zones and pH were assessed. RESULTS: P. micros demonstrated the highest level of in vitro resistance. Pulpdent and Ultracal pastes had the highest pH (12.64 and 12.53, respectively). The addition of Pulpdent to Ledermix did not increase the zone sizes significantly. CONCLUSIONS: All the commercial products showed some in vitro antimicrobial activity. Ledermix paste and the 50:50 Ledermix/Pulpdent mixture being the most effective in this model. The known anti-inflammatory/analgesic properties of Ledermix and the results from this agar model suggest that the 50:50 Ledermix/Pulpdent combination would be the preferred medicament for clinical use in symptomatic cases, even though the addition of calcium hydroxide to Ledermix did not appear to be synergistic in terms of enhancing the antimicrobial action.

Identification of cultivable microorganisms from root canals with apical periodontitis following two-visit endodontic treatment with antibiotics/steroid or calcium hydroxide dressings.

The study was aimed at comparing the efficacy of disinfection of root canals with periapical radiolucencies when treated with either antibiotics/steroid medicaments (Ledermix or Septomixine) or a calcium hydroxide paste (Calasept). Microbiological samples were taken before and after two-visit endodontic treatment from 88 canals with apical periodontitis. All of the canals but one (87 of 88) had cultivable growth before treatment. After dressing with Ledermix, Septomixine, or Calasept, the percentages of canals remained with positive growth were 48% (13 of 27), 31% (8 of 26), and 31% (11 of 35), respectively. The chi(2) tests showed there were no significant differences in the number of canals with positive growth or mean colony forming units counts after instrumentation, irrigation and dressing. In the Ledermix group, 38 strains of bacteria were recovered. The Septomixine group had 25 strains, and the Calasept group had 25 strains. Gram-positive facultative anaerobic cocci (including staphylococci and streptococci) were more prevalent than the Gram-negative obligate anaerobic rods after treatment in all three groups. Similarities in the reduced number of canals with residual growth, and the prevalence of Gram-positive facultative anaerobic cocci suggest that the use of different inter-appointment dressings produced similar microbiological outcomes. However, factors other than the antimicrobial effectiveness of intracanal medicaments may also be responsible for the results observed.

Effects of tetracyclines on aldosterone- and insulin-mediated Na+ transport in the toad urinary bladder.

The effect of oxytetracycline and demethylchlortetracycline on aldosterone- and insulin-mediated Na+ transport (short-circuit current) were examined in toad urinary bladders mounted in modified Ussing chambers. Oxytetracycline had little or no effect on either basal or aldosterone-mediated Na+ transport. In contrast, demethylchlortetracycline markedly inhibited both basal and aldosterone-mediated Na+ transport. Furthermore, demethylchlortetracycline inhibited the aldosterone response significantly out of proportion to its effects on basal Na+ transport. Neither of the drugs had an effect on insulin-mediated Na+ transport. Consequently, the natriuresis observed in certain patients treated with demethylchlortetracyline may be related to drug-induced renal resistance to the effects of aldosterone.

Atrial natriuretic peptide in patients with the syndrome of inappropriate antidiuretic hormone secretion and with diabetes insipidus.

To examine a possible role for atrial natriuretic peptide (ANP) in water and sodium metabolism disturbances associated with abnormal vasopressin (AVP) secretion, we measured plasma ANP concentrations in 15 patients with the syndrome of inappropriate antidiuretic hormone secretion (SIADH) and in 17 patients with central diabetes insipidus (DI). The mean plasma ANP concentration (30.2 +/- 10.4 pmol/L) in SIADH patients who had hyponatremia, plasma hypoosmolality, hyperosmolar urinary compared to plasma sodium levels, and increased plasma AVP levels relative to plasma osmolality was significantly higher than that in normal subjects (12.6 +/- 4.9 pmol/L), although there was a considerable individual variation in plasma ANP ranging from normal to clearly elevated levels (15.1-47.0 pmol/L). When hyponatremia was corrected by water restriction or demeclocycline administration, plasma ANP levels decreased significantly and fell into the normal range (12.5 +/- 4.3 pmol/L). DI patients who complained of polyuria and polydipsia and had hypoosmolar urine, normal or elevated plasma sodium concentrations, and decreased plasma AVP levels relative to plasma osmolality, on the other hand, had a significantly lower mean plasma ANP level (7.6 +/- 2.9 pmol/L) than normal subjects. There was, again, a considerable overlap between plasma ANP levels in individual DI patients (4.2-13.9 pmol/L) and those in normal subjects. Treatment with 1-desamino-8-D-arginine vasopressin resulted in a significant increase in the mean plasma ANP level (18.6 +/- 8.0 pmol/L). There were no significant correlations between plasma ANP and AVP levels in either group of patients. The results indicate that ANP secretion is modulated by changes in plasma volume consequent to abnormal AVP secretion, which may have a pathophysiological significance in maintaining volume homeostasis.

The ocular hypotensive effects of demeclocycline, tetracycline and other tetracycline derivatives.

Demeclocycline, tetracycline and other tetracycline derivatives lowered intraocular pressure (IOP) in rabbits following intravitreal injection, but the onset of this effect was not evident until 1 or more days after drug administration. Of the drugs tested, demeclocycline was the most active ocular hypotensive agent. Demeclocycline caused a dose-dependent decrease in IOP. The maximum IOP decrease of approximately 12 mm Hg occurred 5 days after intravitreal administration of 0.5 mg, with the effect persisting for over a week. Demeclocycline did not alter tonographically measured aqueous humor outflow facility or episcleral venous pressure. Based on calculated aqueous humor flow rates following 0.2 mg demeclocycline, a 62% decrease in aqueous humor formation occurred 7 days after intravitreal injection. The flow-to-diffusion ratio for ascorbate was reduced 54% 6 days after the intravitreal administration of demeclocycline, a change also consistent with suppression of aqueous humor formation. Anterior chamber aqueous humor protein concentration was increased 6 days after demeclocycline administration. No histologic changes were present in the treated eyes by light microscopy. Intravitreal demeclocycline similarly lowered IOP in cats, with the duration of effect lasting up to 20 days.

Inhibition of thrombin-induced secretion from platelets by chlortetracycline and its analogs.

Chlortetracycline (CTC) (1.0 mM) blocks platelet secretion after a few seconds preincubation. The amount needed for inhibition can be reduced relative to time of preincubation. 50 micro M CTC. Two tetracycline analogs, anhydrotetracycline and demeclocycline (DMC), have different solubility properties in nonionic medium, but inhibit secretion at the same concentration, with little effect on the metabolic ATP level. The results suggest that CTC and its analogs do not inhibit platelet function by acting as metabolic inhibitors. While CTC causes increased leakage of cytoplasmic content at the concentrations where secretion is blocked more than 90%, DMC doses not cause leakage even at much higher concentration, so that there seems to be no connection between the induction of leakage (i.e. the membrane-active properties) and the inhibitory effect of the drugs.

Transepithelial water movement in response to carbamazepine, chlorpropamide and demeclocycline in toad urinary bladder.

1. Osmotic water movement across toad isolated hemibladders was measured by a gravimetric method. 2. The influence of carbamazepine, chlorpropamide and demeclocycline on the antidiuretic hormone (ADH)-induced water flow rate was examined. 3. No antidiuretic activity due to carbamazepine alone was observed but a slight inhibition due to ADH-induced water flow was observed in the presence of carbamazepine over a selected dose-range. This was unexpected and is inconsistent with data from in vivo studies in man. 4. Chlorpropamide potentiated ADH-induced water flow, in keeping with the hypothesis that chlorpropamide sensitizes the renal tubules to ADH-induced water flow. 5. Demeclocycline inhibited ADH-induced water flow. The mechanism of action remains unclear.

Sensitivity of Brucella spp to tetracycline and its analogues.

The sensitivity to eight tetracyclines of 100 strains of brucellae, comprising strains of Brucella abortus, Br. melitensis, and Br. suis, was determined. Demethylchlortetracycline was the most effecitve against all the groups of brucellae, whereas oxytetracycline and chlortetracycline were the least effective. The mean MIC value for demethylchlortetracycline, doxycycline, lymecycline, and tetracycline was less than or equal to 1 mug/ml. Strains of Br. abortus biotype 2 and Br. suis were the most sensitive strains examined.

The effect of minocycline on Candida albicans.

Minocycline, a new tetracycline derivative, was found to inhibit the growth of Candida albicans. Inhibition was much affected by the composition of the medium and was difficult to demonstrate in fluid cultures. Study of the rate of budding in shallow broth cultures in Petri dishes showed that the addition of 20 mug/ml minocycline prolonged the lag phase by three hours. C. tropicalis was similarly inhibited and C. guilliermondii and C. parapsilosis to a lesser degree. Six other tetracyclines were tested and found to inhibit Candida only in very high concentrations.

Anthracene with near ultraviolet light inhibiting epidermal proliferation.

Anthracene plus near ultraviolet (UV) light (UV-A, 320 to 400 nm) suppresses DNA synthesis and mitosis in mouse epidermis. Ultraviolet-A light or anthracene alone does not have any effect. There is no photoactivation of anthracene to enhance depression of DNA synthesis by either UV-B (290 to 320 nm) or UV-C (254 nm) light. While methoxsalen with UV-A light inhibits DNA synthesis, the phototoxic drugs chlorpromazine hydrochloride and demethylchlortetracycline do not. The combination of anthracene plus UV-A light may have therapeutic effectiveness for psoriasis with less potential for photocarcinogenesis than psoralens plus UV-A light.

A comparative study on the effects of tetracyclines and lithium on the cyclic AMP second messenger system in rat brain.

1. This study was aimed at investigating the effects of demeclocycline (DMC), minocycline (MC), and lithium (Li) in vitro on cyclic AMP (cAMP) accumulation in rat cerebral cortex stimulated by noradrenaline, forskolin, and ouabain. 2. DMC, MC, and Li dose-dependently reduced noradrenaline-stimulated cAMP formation in cortical slices, but only Li inhibited the cAMP formation induced by forskolin. 3. In contrast to Li, DMC and MC did not affect noradrenaline-stimulated adenylate cyclase activity in cortical membranes. 4. In cortical slices, ouabain stimulated the cAMP production (required the presence of extracellular Ca2+ and was blocked by verapamil). Ouabain-stimulated cAMP accumulation in cortical slices was inhibited by DMC, MC, and Li. 5. DMC and MC do not seem to interact directly with the adenylate cyclase as reported for Li. It is concluded that the tetracyclines, DMC and MC, affect the cAMP signaling system in rat brain by mechanisms that differ from that of Li. The decreased receptor agonist-stimulated cAMP production in cortical slices in the presence of DMC and MC may be due to the Ca(2+)-chelating ability of these tetracyclines.

Effects of repeated oral doses of demethylchlortetracycline on bones and dentin of young rhesus monkeys.

Seven oral doses of demethylchlortetracycline were administered to monkeys that received serial injections of lead acetate to intravitally stain calcification sites. Bone growth was greatly inhibited, whereas dentin apposition was spared from the cumulative toxicity of demethylchlortetracycline. Cessation of bone growth and its duration could be correlated with serum levels.

Effects of demethylchlortetracycline phototoxicity on the structure and functions of rat liver mitochondria.

The phototoxic effects of demethylchlortetracycline (DMCT) with UVA radiation on isolated rat liver mitochondria were studied. DMCT at concentration of 21.5 microM with 5.53 x 10(-2) J cm-3 of UVA was found to be a potent uncoupler of oxidative phosphorylation. DMCT alone also uncoupled mitochondria but at higher concentrations (105 microM). ATPase activity was remarkably induced in mitochondria exposed to DMCT (21.5 microM) plus UVA (120% of DNP-stimulated ATPase activity). Content of ATP in such mitochondria when measured after addition of ADP was much smaller than that in control mitochondria. Ultrastructurally, mitochondria treated either with DMCT (21.5 microM) or with UVA alone stayed in the condensed configuration. On the other hand, uncoupled mitochondria treated with DMCT plus UVA became swollen and were changed into the orthodox configuration.

Inhibition of E-and EAC-rosette formation by gentamicin, ampicillin and tetracycline.

The determination of the absolute number of cells with receptors for sheep erythrocytes and for C3 by rosette techniques are usual clinical tests in the evaluation of the lymphoid populations in different diseases. Antibiotics are commonly added to avoid contamination in the reagents employed in the rosetting studies, as well as in "in vitro" tests of lymphocytes function and in cultures of cell lines. In this study we have established the influence of gentamicin (G), demeclocycline hydrochloride (DH) and ampicillin (A) on the membrane receptors, through their effect on E4 degrees C and EAC formation. We found that in the presence of each of the three antibiotics there was a partial inhibition of both rosettes. The inhibition was found to be dose related, gentamicin concentrations of 85 micrograms/ml and 42,5 micrograms/ml resulting in significant reduction of E4 degrees C and EAC rosettes respectively. Studies with A and DH indicated that EAC rosettes were inhibited by both drugs after incubation times of at least 15 minutes, while a minimum of 30 minutes of incubation was required to attain significant inhibition of E4 degrees C rosettes. These results indicate that the C3 receptors would be more sensitive to the action of these antibiotics than the receptors for sheep erythrocytes.

Comparative effects of intrauterine instillation of analogues of quinacrine and tetracycline on uterine morphology in the rat.

Currently, intrauterine instillation of quinacrine hydrochloride is used to induce closure of the uterotubal junction in women, thus constituting a chemical method of sterilization. Questions regarding the safety of this drug have been raised. The purpose of the present study is to screen other drugs for their sterilizing potential by comparing quinacrine-induced changes in uterine morphology in the rat which have previously been correlated with decreased fertility with the changes induced by other drugs. The drugs tested include quinacrine-like compounds, namely chloroquine, primaquine and trimethoprim; and tetracycline and its analogues which are known sclerosing agents. The quinacrine-like drugs were relatively ineffective in producing uterine lesions similar to those of quinacrine, but like quinacrine, chloroquine and primaquine showed some toxicity. Tetracycline and its analogues produced quinacrine-like morphologic changes in the rat uterus and showed no toxicity for the doses tested. These results prompt further testing of tetracycline and its analogues as sterilizing agents.

Effect of intrauterine administration of tetracyclines on cynomolgus monkeys.

Cynomolgus monkeys were used to screen for chemicals which potentially could be used as tubal occluding agents. Intrauterine administrations of solution or pellets of tetracycline and its analogues (100 mg doses) were tested for their effects on morphologic changes in the reproductive tract of monkeys. These effects were compared to monkeys receiving intrauterine administration of quinacrine pellets (36 mg) since quinacrine has been used successfully in the clinical setting. Blood levels of drugs, blood chemistry and hematology determinations and liver and kidney pathology data were also obtained as indices for toxicity. Morphologic damage to the uterine lining and intramural section of the tube (including necrosis, inflammation or scarring) was elicited by intrauterine tetracycline and doxycycline in the same frequency and severity as quinacrine. In contrast, saline or sham control monkeys showed no morphological damage of the tube or uterus. Although all drugs could be detected in the blood 4 hours after intrauterine administration, levels were near or below the limit of detection by one week. No evidence was found for toxicity of tetracycline or its analogues for the dosage given. Because of these results and the extensive literature on tetracycline toxicity, further studies should be directed toward the use of tetracycline as a sterilizing agent in women.

Inhibition by antibiotic tetracyclines of rat cortical noradrenergic adenylate cyclase and amphetamine-induced hyperactivity.

Two antibiotic tetracyclines, demeclocycline (DMC) and minocycline, share several biochemical and behavioral properties with lithium (Li). DMC inhibited both noradrenaline- and chloradenosine-sensitive cyclic AMP accumulation in rat cerebral cortical slices both in vitro and ex vivo following two weeks of chronic dietary treatment. Minocycline, a lipophilic tetracycline, produced similar results in vitro. Both DMC and minocycline reduced open-field activity levels in rats following acute treatment, four hours prior to testing. Moreover, both drugs inhibited amphetamine-induced hyperactivity in the open field. Chronic treatment with 0.4% and 0.8% dietary DMC for two weeks attenuated amphetamine hyperactivity without affecting baseline activity levels in the open field. Neither DMC nor minocycline attenuated apomorphine-induced stereotypy at doses that attenuated amphetamine hyperactivity, a profile which is similar to that of lithium. Unlike lithium, however, DMC did not reverse reserpine-induced hypoactivity.

Effect of calcium hydroxide and combinations of Ledermix and calcium hydroxide on inflamed pulp in dog teeth.

The effects of Ledermix+calcium hydroxide (Ca(OH)2) or Ca(OH)2 alone on inflamed pulp tissues of dogs were studied. Fifty-nine upper incisor teeth of 10 dogs were used. Class V cavities were prepared and filled with amalgam after placement of decayed dentin particles. After 7 days, the decayed dentin and alloy were removed, and the pulps of the teeth were exposed. Ledermix and Ca(OH)2 mixtures or Ca(OH)2 alone were applied to the cavities. At the end of 7, 30, and 90 days, the teeth were extracted and examined histopathologically. Inflammation was found to be more prevalent in the 7- and 30-day groups that were treated with the Ledermix+Ca(OH)2 combination, whereas fibrosis and necrosis were nearly similar in both groups. In the 90-day groups, no inflammation was seen. No difference between the two 90-day groups with regard to reparative dentin was found.