Re-exposure Period Research Articles

Relapse-like behavior in a mouse model of the OPRM1 (mu-opioid receptor) A118G polymorphism: Examination with intravenous oxycodone self-administration

The widely abused prescription opioid oxycodone is a mu-opioid receptor (MOP-r) agonist and addiction to such opioids is a relapsing disorder. The human MOP-r gene (OPRM1) has an important functional single nucleotide polymorphism (SNP), A118G, which affects risk of severe opioid use disorders. A112G (G/G) knock-in mice are models of human A118G carriers. We examined oxycodone self-administration (SA) in male and female G/G versus wild type (A/A) mice in SA sessions and in relapse-like behavior. Adult male and female G/G and A/A mice self-administered oxycodone (0.25 mg/kg/infusion, FR1) for 10 consecutive days. Following 10-day home cage drug free withdrawal, the mice were re-exposed to oxycodone SA for a further 10 days. MOP-r receptor mRNA in various brain regions were examined immediately after the last re-exposure session.We found that G/G mice had greater oxycodone SA than A/A mice in the initial and in re-exposure sessions. Mice of both genotypes had greater oxycodone intake during the re-exposure period than during the initial exposure. We also detected differences in MOP-r gene expression due to genotype, sex and oxycodone SA history in the dorsal striatum, hippocampus, and prefrontal cortex. These studies may improve our understanding of MOP-r-agonist self-exposure and relapse in human carriers of the A118G SNP.

Cessation of fluoxetine treatment increases alcohol seeking during relapse and dysregulates endocannabinoid and glutamatergic signaling in the central amygdala.

Administration of selective serotonin reuptake inhibitors (SSRIs), typically used as antidepressants, induces long-lasting behavioral changes associated with alcohol use disorder (AUD). However, the contribution of SSRI (fluoxetine)-induced alterations in neurobiological processes underlying alcohol relapse such as endocannabinoid and glutamate signaling in the central amygdala (CeA) remains largely unknown. We utilized an integrative approach to study the effects of repeated fluoxetine administration during abstinence on ethanol drinking. Gene expression and biochemical and electrophysiological studies explored the hypothesis that dysregulation in glutamatergic and endocannabinoid mechanisms in the CeA underlie the susceptibility to alcohol relapse. Cessation of daily treatment with fluoxetine (10 mg/kg) during abstinence resulted in a marked increase in ethanol seeking during re-exposure periods. The increase in ethanol self-administration was associated with (a) reductions in levels of the endocannabinoids N-arachidonoylethanolomine and 2-arachidonoylglycerol in the CeA, (b) increased amygdalar gene expression of cannabinoid type-1 receptor (CB1), N-acyl phosphatidylethanolamine phospholipase D (Nape-pld), fatty acid amid hydrolase (Faah), (c) decreased amygdalar gene expression of ionotropic AMPA (GluA2 and GluA4) and metabotropic (mGlu3) glutamate receptors, and (d) increased glutamatergic receptor function. Overall, our data suggest that the administration of the antidepressant fluoxetine during abstinence dysregulates endocannabinoid signaling and glutamatergic receptor function in the amygdala, facts that likely facilitate alcohol drinking behavior during relapse.

Correlation between antibiotic-induced feeding depression and body size reduction in zooplankton (rotifer, Brachionus calyciflorus): Neural response and digestive enzyme inhibition

The study analyzed the correlation between the antibiotic-induced feeding depression and body size reduction in rotifer, Brachionus calyciflorus, involving exposure, post-exposure and re-exposure periods. The filtration and ingestion rates of the rotifers were inhibited in these three exposure periods at any given concentration of the antibiotic sulfamethazine (SMZ). As food for rotifer, the cell size of the green algae was unchanged, which indicated that it could not drive feeding depression. Secondly, several corresponding physiological responses were considered. Reactive oxygen species (ROS) levels increased in the post-exposure and the re-exposure; acetylcholinesterase (AChE) activity was significantly decreased in the exposure and the re-exposure, whereas it was induced in the post-exposure. The activities of amylase and lipase were always inhibited in these three exposure periods. Additionally, significant decreases in lorica length, width and biovolume of rotifers occurred after the feeding depression. Statistical analysis indicated a positive correlation between the activity of the digestive enzyme and the body size. Our results demonstrated that SMZ could influence the neurotransmission, inhibit the activity of the digestive enzyme, and finally result in body size reduction. These results provided an integrated perspective on assessing the toxicity effects of antibiotic in non-lethal dosage on the feeding behavior of non-target aquatic organisms.

Acute and chronic modulation of striatal endocannabinoid-mediated plasticity by nicotine.

The endocannabinoid (eCB) system modulates several phenomena related to addictive behaviors, and drug‐induced changes in eCB signaling have been postulated to be important mediators of physiological and pathological reward‐related synaptic plasticity. Here, we studied eCB‐mediated long‐term depression (eCB‐LTD) in the dorsolateral striatum, a brain region critical for acquisition of habitual and automatic behavior. We report that nicotine differentially affects ex vivo eCB signaling depending on previous exposure in vivo. In the nicotine‐naïve brain, nicotine facilitates eCB‐signaling and LTD, whereas tolerance develops to this facilitating effect after subchronic exposure in vivo. In the end, a progressive impairment of eCB‐induced LTD is established after protracted withdrawal from nicotine. Endocannabinoid‐LTD is reinstated 6 months after the last drug injection, but a brief period of nicotine re‐exposure is sufficient to yet again impair eCB‐signaling. LTD induced by the cannabinoid 1 receptor agonist WIN55,212‐2 is not affected, suggesting that nicotine modulates eCB production or release. Nicotine‐induced facilitation of eCB‐LTD is occluded by the dopamine D2 receptor agonist quinpirole, and by the muscarinic acetylcholine receptor antagonist scopolamine. In addition, the same compounds restore eCB‐LTD during protracted withdrawal. Nicotine may thus modulate eCB‐signaling by affecting dopaminergic and cholinergic neurotransmission in a long‐lasting manner. Overall, the data presented here suggest that nicotine facilitates eCB‐LTD in the initial phase, which putatively could promote neurophysiological and behavioral adaptations to the drug. Protracted withdrawal, however, impairs eCB‐LTD, which may influence or affect the ability to maintain cessation.

Integrated toxic evaluation of sulfamethazine on zebrafish: Including two lifespan stages (embryo-larval and adult) and three exposure periods (exposure, post-exposure and re-exposure)

Persistence of antibiotics in aquatic environment may pose a risk to the non-target aquatic organisms. This study provided an integrated evaluation to analyze the toxic stress of sulfamethazine (SMZ) on zebrafish in two lifespan stages (embryo-larval and adult) and three exposure periods (exposure, post-exposure and re-exposure). Zebrafish embryos and adult zebrafish were exposed to SMZ at 0.2, 20 and 2000 μg/L, respectively. The results showed that SMZ at any given concentration inhibited the hatching of embryos at 58–96 hpf (hours post-fertilization). Our result also indicated that two major kinds of the malformation, which was induced by the antibiotic, were edema and spinal curvature. Additionally, the antibiotic stimulated the heartbeat while reduced the body length of the embryo at 72 hpf. Superoxide dismutase (SOD) activities and malondialdehyde (MDA) contents significantly increased at 120 hpf when the embryos were exposed to the lowest concentration (0.2 μg/L) of the antibiotic. On the other hand, the antibiotic induced SOD activities and MDA contents in adult zebrafish in the exposure and re-exposure periods. The MDA contents could recover while SOD activities still increased in 2 d after the exposure. Both SOD activities and MDA contents could recover in 7 d after the exposure. Levels of SOD and MDA in the re-exposure were higher than those in the first exposure. Our results suggested that SMZ had toxic effects on both embryos and adult zebrafish, and provided an integrated evaluation of the toxic effects of SMZ on zebrafish at a new perspective.

The impact assessment of anticancer drug imatinib on the feeding behavior of rotifers with an integrated perspective: Exposure, post-exposure and re-exposure

The anticancer drugs are getting increasing attention as an emerging contaminant in the aquatic environments. In the present study, feeding behavior of the rotifer Brachionus calyciflorus under the impact of anticancer drug imatinib was evaluated. Traditional toxicological studies usually focus on dose-effect relationship at a given exposure time, while ignore the possible impact after the exposure. Thus, how the impact varied in the post-exposure and re-exposure was also considered in the present study. The feeding depression of the rotifers was attributed to the increased concentration of imatinib. Although the filtration and ingestion rate of the rotifers recovered to a certain extent after the exposure, the significant feeding inhibition still persisted even if the exposure was ended. In the re-exposure period, the feeding behavior was less depressed than those of the exposure period, which implied that rotifers might develop a tolerance to the same toxics. The activities of acetylcholine esterase (AchE) and the levels of reactive oxygen species (ROS) in rotifers were also detected. Imatinib inhibited the activities of AchE in the exposure and re-exposure while ROS levels increased significantly in the re-exposure period. Our present study provided an integrated assessment the potential environmental risks of imatinib at a new perspective.

Ethanol Sensitization during Adolescence or Adulthood Induces Different Patterns of Ethanol Consumption without Affecting Ethanol Metabolism.

In previous study, we demonstrated that ethanol preexposure may increase ethanol consumption in both adolescent and adult mice, in a two-bottle choice model. We now questioned if ethanol exposure during adolescence results in changes of consumption pattern using a three-bottle choice procedure, considering drinking-in-the-dark and alcohol deprivation effect as strategies for ethanol consumption escalation. We also analyzed aldehyde dehydrogenase (ALDH) activity as a measurement of ethanol metabolism. Adolescent and adult Swiss mice were treated with saline (SAL) or 2.0 g/kg ethanol (EtOH) during 15 days (groups: Adolescent-SAL, Adolescent-EtOH, Adult-SAL and Adult-EtOH). Five days after the last injection, mice were exposed to the three-bottle choice protocol using sucrose fading procedure (4% + sucrose vs. 8%–15% ethanol + sucrose vs. water + sucrose) for 2 h during the dark phase. Sucrose was faded out from 8% to 0%. The protocol was composed of a 6-week acquisition period, followed by four withdrawals and reexposures. Both adolescent and adult mice exhibited ethanol behavioral sensitization, although the magnitude of sensitization in adolescents was lower than in adults. Adolescent-EtOH displayed an escalation of 4% ethanol consumption during acquisition that was not observed in Adult-EtOH. Moreover, Adult-EtOH consumed less 4% ethanol throughout all the experiment and less 15% ethanol in the last reexposure period than Adolescent-EtOH. ALDH activity varied with age, in which older mice showed higher ALDH than younger ones. Ethanol pretreatment or the pattern of consumption did not have influence on ALDH activity. Our data suggest that ethanol pretreatment during adolescence but not adulthood may influence the pattern of ethanol consumption toward an escalation in ethanol consumption at low dose, without exerting an impact on ALDH activity.

Multigenerational effect of perfluorooctane sulfonate (PFOS) on the individual fitness and population growth of Daphnia magna

We investigated the multigenerational effect of PFOS to individual fitness (e.g., body weight, acetylcholinesterase and glutathione S-transferase) and population growth (e.g., offspring number and time to first brood) of Daphnia magna during continuous and discontinuous exposures. The intrinsic rate of population growth was also calculated. In the continuous exposure, population growth-related adverse effects were detected during all test periods, and the adverse effect tended to be weaker in later generations. On the other hand, individual fitness-related adverse effects were observed from F1 not in F0 and deteriorated as the generation number increased. These results imply that individual fitness worsens although the population growth is restored in later generations. Upon discontinuous exposure, a few but significant adverse effects were observed during the non-exposure period and highest effects were detected during the re-exposure period. This encourages the study of different exposure scenarios, which may result in unexpected and higher PFOS toxicity. Consequently, this study confirms adverse effects of PFOS to Daphnia magna in multigenerational period and supports reasons for studies linking individual fitness changes to population dynamics and covering diverse exposure scenarios to evaluate the risk of PFOS in a water environment.

Changes in the incentive value of food after naltrexone treatment depend on a differential preference for a palatable food in male rats

Introduction: Opioid antagonist treatments such as naltrexone (NTX) and naloxone reduce consummatory behavior of palatable food (PF) and other incentives. Meanwhile, a significant increase in alcohol consumption has been observed when it is offered immediately after ending NTX treatment, an effect apparently produced by increased opioidergic activity caused by up-regulation of opioid receptors.Objective: On this basis we assessed changes in the consumption of PF after opioid antagonist treatment in rats with different preferences for that food.Methods: The preference of male Wistar rats for a PF was classified as low, medium, or high in a baseline period, after which the animals in each preference level were sub-divided into control and experimental groups that received injections of either NTX (2 mg/kg twice per day/6 days) or a saline solution with a space of 8 hours between doses. At the end of pharmacological treatment (PT), subjects were re-exposed to the PF.Results: Increased PF intake was found only in the low-preference group, but the increase was observed in both the experimental and control animals. Also, a decrease in chow intake during PT was observed in all preference groups, while recovery after treatment was noted only in the low-preference rats.Discussion: The increased intake observed in the low-preference rats after treatment phase suggests that the hypothalamic–pituitary–adrenal axis activation during PT could have enhanced the rewarding characteristics of the sweet food and so facilitated and increased its consumption in the re-exposure period.

Organized breast screening improves reattendance compared to physician referral: a case control study.

BackgroundThe Ontario Breast Screening Program (OBSP) is a population-based breast screening programme, not requiring physician referral. OBSP invites women by mail to book their next screens. However, women who do not participate in the OBSP, may be referred by physicians to non-OBSP mammography facilities, which do not remind women to book their next screen.MethodsWe identified women without breast cancer prior to June 30, 2011, having bilateral mammography (M) during a baseline period at age 50 – 69 at OBSP or non-OBSP facilities, and during a re-exposure period, at the same facility type. We used a case-control design to study the association of facility type and having M during an outcome period. Cases were women failing to receive the outcome M. Controls were matched by age, census tract, and socioeconomic status. Exposure was baseline facility type. Covariates were comorbidity, residential mobility, and primary care physician (PCP) characteristics. Conditional logistic regression analysis was performed.ResultsCases were less likely to have been screened at OBSP facilities. Failure to receive the outcome M was associated with having moved after re-exposure M (OR = 1.61, 95% confidence interval (CI) 1.52, 1.71), having a male PCP (OR = 1.05, 95% CI 1.02, 1.05), or a higher Charlson score (OR = 1.06 per unit increase, 95% CI 1.03, 1.09). Having re-exposure M at an OBSP facility (OR = 0.18, 95% CI 0.18, 0.19)., having a Canadian trained PCP (OR = 0.83, 95% CI 0.8, 0.87), and having a PCP one year after the re-exposure M (OR = 0.81, 95% CI 0.68, 0.97) were protective against failure to receive the outcome M.ConclusionsThe OBSP, not requiring physician referral, and inviting women by mail to book their next screen, is associated with a lower probability of failure to reattend for subsequent screening than screening by PCP referral to non-OBSP facilities.

Tolerance and sensitization to chronic escalating dose heroin following extended withdrawal in Fischer rats: possible role of mu-opioid receptors

Heroin addiction is characterized by recurrent cycles of drug use, abstinence, and relapse. It is likely that neurobiological changes during chronic heroin exposure persist across withdrawal and impact behavioral responses to re-exposure. We hypothesized that, after extended withdrawal, heroin-withdrawn rats would express behavioral tolerance and/or sensitization in response to heroin re-exposure and that these responses might be associated with altered mu-opioid receptor (MOPr) activity. Male Fischer rats were exposed chronically to escalating doses of heroin (7.5-75 mg/kg/day), experienced acute spontaneous withdrawal and extended (10-day) abstinence, and were re-exposed chronically to heroin. Homecage behaviors and locomotor activity in response to heroin, as well as somatic withdrawal signs, were recorded. Separate groups of rats were sacrificed after extended abstinence and MOPr expression and G-protein coupling were analyzed using [(3)H]DAMGO and [(35)S]GTPγS assays. The depth of behavioral stupor was lower during the initial days of heroin re-exposure compared to the initial days of the first exposure period. Behavioral responses (e.g., stereotypy) and locomotion were elevated in response to heroin re-exposure at low doses. Rats conditioned for heroin place preference during the chronic re-exposure period expressed heroin preference during acute withdrawal; this preference was stronger than rats conditioned during chronic heroin exposure that followed chronic saline and injection-free periods. Extended withdrawal was associated with increased MOPr expression in the caudate-putamen and frontal and cingulate cortices. No changes in G-protein coupling were identified. Aspects of tolerance/sensitization to heroin are present even after extended abstinence and may be associated with altered MOPr density.

The effects of prolonged darkness on temperate and tropical marine phytoplankton, and their implications for ballast water risk management

Phytoplankton assemblages from tropical (Goa) and temperate (UK) locations were exposed to a 28day dark period, followed by a period of re-exposure to light. During this time phytoplankton survival and changes in nutrient concentrations were mapped. The tropical plankton water samples showed high nutrient levels after the dark period which were utilised by cells during the re-exposure period. UK experiments looked at the effect of three different water types on population recovery after the 28day dark period, and differences due to seasonal effects.The population growth observed during the re-exposure period in the tropical population was comparable to that of the temperate population. Water type affected recovery and of the three tested media fresh seawater promoted the highest levels of growth. Seasonality had a significant influence on species survival. Understanding the effects of all these factors can aid the development of effective risk assessments in ballast water management.

Pandemic (H1N1) 2009 Reinfection, Chile

To the Editor: Since March 2009, influenza A pandemic (H1N1) 2009 has spread worldwide (1), and in South America, Chile was 1 of the countries most affected by the pandemic, with 21.4 cases among every 1,000 persons. Treatment guidelines in Chile recommended antiviral drug treatment with oseltamivir or zanamivir for 5 days for all patients with confirmed or suspected virus subtype H1N1 infection (2). In persons with seasonal influenza, specific antibody responses reach peak titers by 4 weeks after infection and confer protection against the infecting strain and closely related strains (3). Reinfection is rarely seen in nonpandemic influenza A. We report on 3 patients with confirmed influenza A pandemic (H1N1) 2009 reinfection after successful treatment with oseltamivir. Patient 1, a healthy 14-year-old girl, had fever, sore throat, and nasal congestion on clinical examination. Pandemic (H1N1) 2009 infection was diagnosed by viral culture and confirmed by PCR specific for subtype H1N1 (LightMix Kit Inf A swine H1; TIB MOLBIOL, Berlin, Germany, for Roche Diagnostic, Indianapolis, IN, Light Cycler 2.0 instrument). The patient received oseltamivir, and symptoms resolved 48 hours after treatment. Twenty days later, fever, muscle aches, and vomiting developed in the patient. Influenza A virus was isolated by viral culture. The patient received a preliminary diagnosis of seasonal influenza and was treated with amantadine. She recovered from the infection before PCR results confirmed it was caused by pandemic (H1N1) 2009 virus. Patient 2, a 62-year-old woman, experienced a high fever, cough, and nasal congestion during a prolonged hospitalization for bowel resection after intestinal ischemia. Pandemic (H1N1) 2009 was confirmed by PCR and viral culture. Oseltamivir was administered 5 days after the onset of symptoms, and the symptoms resolved within the following 5 days. The patient had a new episode of fever, productive cough, and bronchial obstruction 2 weeks later while still hospitalized. Culture results were positive for influenza, and PCR results were positive for pandemic (H1N1) 2009. The patient was again treated with oseltamivir, and PCR results were negative for influenza after 48 hours of antiviral treatment. Patient 3, a previously healthy 38-year-old man, underwent mitral and aortic valve replacement while hospitalized for acute mitral and aortic endocarditis due to Staphylococcus aureus. Eleven days after surgery, he had a sore throat, nasal congestion, cough, and low-grade fever. PCR test results were positive for pandemic (H1N1) 2009. The patient received oseltamivir, and respiratory symptoms resolved within 5 days. He was discharged from the hospital but was readmitted 18 days later with nasal congestion, cough, and high fever. PCR results were again positive for pandemic (H1N1) 2009, and the patient was successfully treated with oseltamivir. Patient 2 and probably patient 3 acquired their infections while hospitalized, suggesting potential nosocomial transmission. No other respiratory viruses were found in any of these patients. The viral isolates were all tested (LightMix for detection of influenza A virus oseltamivir resistance [H274Y]; TIB MOLBIOL) for possible resistance to oseltamivir, but none had the resistance-implicated H274Y mutation in the neuraminidase gene Shedding of seasonal influenza A virus ceases within 5–7 days during natural infection and during infections treated with neuraminidase inhibitors (4). Although clearing of virus after the first infection was not documented in the 3 patients described here, it is unlikely that virus persisted between the 2 episodes of influenza since each of the patients fully recovered after specific antiviral drug treatment. However, we cannot rule out that patient 2 may have never cleared the virus due to her immune suppression. As described by mathematical modeling (5), the 3 patients described were susceptible to reinfection with pandemic (H1N1) 2009 due to the high rate of community infection and to their incomplete immunologic protection within the period of reexposure. During the current pandemic of influenza subtype H1N1, healthcare workers and patients should be aware that symptomatic reinfection might occur after a first episode of infection.

Early malnutrition favours heavy ethanol intake in weaning rats without long-lasting effects

Introduction: Although food restriction is well known to increase ethanol intake, the subject has not been extensively studied in developing animals which could be more vulnerable to long-lasting effects. Therefore, the aim of this study was to show some findings concerning this subject.Materials and methods: Food restriction was used to produce malnutrition either during lactation (lactating dams food restricted 60%) or after weaning (pups food restricted 60%). At weaning, day 25, males were assigned to one of the following groups: C, food ad libitum throughout the experiment – control group; MW, malnourished only after weaning; ML, malnourished only during lactation period; and MLW, malnourished throughout the experiment, during lactation and after weaning. All rats were kept isolated in cages in which they could choose to drink either a 10% ethanol solution or tap water (from days 25 to 45). Re-exposure to this model was performed on day 49. Between exposure and re-exposure, rats drank tap water for 4 days.Results: There was a significant effect of malnutrition during lactation, up to day 35, with heavy drinking patterns (ethanol intake day 2: C, 8 g/kg; MW, 9 g/kg; ML, 19 g/kg; and MLW, 22 g/kg). This heavy drinking pattern lasted until the recovery of body weight. Food restriction after weaning had significant effects after 14 days, when a statistically significant decrease in body weight occurred (body weight day 39: C, 147.8 g; MW, 98.5 g). Only rats which were persistently malnourished (MLW and MW) drank ethanol more significantly than their ad libitum-fed counterparts during the re-exposure period (ethanol intake: malnourished, 5 g/kg; and well-nourished, 2.5 g/kg). Adulteration of the ethanol solution with quinine (0.1 g/l) precluded the effect of malnutrition.Conclusions: Malnutrition during early development had no long-lasting effects on ethanol consumption. In addition, malnutrition increased ethanol consumption as long as it kept body weight low, which was apparently more significant in young animals.

PRECLINICAL STUDY: Effects of concurrent access to multiple ethanol concentrations and repeated deprivations on alcohol intake of high‐alcohol‐drinking (HAD) rats

High-alcohol-drinking rats, given access to 10% ethanol, expressed an alcohol deprivation effect (ADE) only after multiple deprivations. In alcohol-preferring (P) rats, concurrent access to multiple ethanol concentrations combined with repeated cycles of EtOH access and deprivation produced excessive ethanol drinking. The current study was undertaken to examine the effects of repeated alcohol deprivations with concurrent access to multiple concentrations of ethanol on ethanol intake of HAD replicate lines of rats. HAD-1 and HAD-2 rats received access to 10, 20 and 30% (v/v) ethanol for 6 weeks. Rats from each replicate line were assigned to: (1) a non-deprived group; (2) a group initially deprived of ethanol for 2 weeks; or (3) a group initially deprived for 8 weeks. Following the restoration of the ethanol solutions, cycle of 2 weeks of ethanol exposure and 2 weeks of alcohol deprivation was repeated three times for a total of four deprivations. Following the initial ethanol deprivation period, deprived groups significantly increased ethanol intakes during the initial 24-hour re-exposure period. Multiple deprivations increased ethanol intakes, shifted preference to higher ethanol concentrations and prolonged the duration of the elevated ethanol intakes for up to 5 days. In addition, repeated deprivations increased ethanol intake in the first 2-hour re-exposure period as high as 5-7 g/kg (which are equivalent to amounts consumed in 24 hours by HAD rats), and produced blood ethanol levels in excess of 150 mg%. The results indicate that HAD rats exhibit 'loss-of-control' of alcohol drinking with repeated deprivations when multiple ethanol concentrations are available.

Caries-Preventive Effect of a One-Time Application of Composite Resin and Glass Ionomer Sealants after 5 Years

The aim of the present trial was to (1) compare the caries-preventive effect of glass ionomer sealants, placed according to the atraumatic restorative treatment (ART) procedure, with composite resin sealants over time and (2) investigate the caries-preventive effect after complete disappearance of sealant material. Forty-six boys and 57 girls, mean age 7.8 years, were randomly divided into two treatment groups in a parallel-group study design. A light-polymerized composite resin sealant material and a high-viscosity glass ionomer were each placed in 180 fully erupted first molars in their respective treatment groups. Evaluation took place annually for 5 years by calibrated examiners. After 5 years, 86% composite resin and 88% glass ionomer sealants did not survive. Three categories of re-exposure periods for caries development in pits and fissures after complete loss of sealants were distinguished: 0–1, 1–2 and 2–3 years. In the 2- to 3-year group, 13 and 3% of pits and fissures previously sealed with composite resin and glass ionomer, respectively, were diagnosed as having developed a dentine lesion. The relative risks (95% CI) of dentine lesion development in surfaces sealed with glass ionomer compared to those sealed with composite resin after 3, 4 and 5 years were 0.22 (0.06–0.82), 0.32 (0.14–0.73) and 0.28 (0.13–0.61), respectively. The relative risks of dentine lesion development in pits and fissures previously sealed with glass ionomer compared with composite resin over re-exposure periods of 1–2 and 2–3 years were 0.26 (0.14–0.48) and 0.25 (0.09–0.68), respectively. We conclude that the caries-preventive effect of high-viscosity glass ionomer sealants, placed using the ART procedure, was between 3.1 and 4.5 times higher than that of composite resin sealants after 3–5 years. Furthermore, high-viscosity (ART) glass ionomer sealants appear to have a four times higher chance of preventing caries development in re-exposed pits and fissures of occlusal surfaces in first molars than light-cured composite resin sealant material over a 1- to 3-year period. A well-designed clinical trial using different types of oral health personnel should be implemented to confirm these initial results.

Effects of Long‐Term Episodic Access to Ethanol on the Expression of an Alcohol Deprivation Effect in Low Alcohol–Consuming Rats

The alcohol-preferring (P) and -nonpreferring (NP) and high alcohol-drinking (HAD) and low alcohol-drinking (LAD) rats have been selectively bred for divergent preference for ethanol over water. In addition, both P and HAD rats display an alcohol deprivation effect (ADE). This study was undertaken to test whether the NP, LAD-1, and LAD-2 lines of rats could display an ADE as well. Adult female NP, LAD-1, and LAD-2 rats were given concurrent access to multiple concentrations of ethanol [5, 10, 15% (v/v)] and water in an ADE paradigm involving an initial 6 weeks of 24-hr access to ethanol, followed by four cycles of 2 weeks of deprivation from and 2 weeks of re-exposure to ethanol (5, 10, and 15%). A control group had continuous access to the ethanol concentrations (5, 10, and 15%) and water through the end of the fourth re-exposure period. For NP rats, a preference for the highest ethanol concentration (15%) was evident by the end of the fifth week of access (approximately 60% of total ethanol fluid intake). Contrarily, LAD rats did not display a marked preference for any one concentration of ethanol. All three lines displayed an ADE after repeated cycles of re-exposure to ethanol, with the general ranking of intake being LAD-1 > NP > LAD-2 (e.g., for the first day of reinstatement of the third re-exposure cycle, intakes were 6.5, 2.9, and 2.4 g/kg/day compared with baseline values of 3.1, 2.0, and 1.3 g/kg/day for each line, respectively). By the 13th week, rats from all three lines, with a ranking of LAD-1 > NP > LAD-2, were drinking more ethanol (3.3, 2.2, and 2.0 g/kg/day, respectively) compared with their consumption during the first week of access (approximately 1.1 g/kg/day for all three lines). These data indicate that access to multiple concentrations of ethanol and exposure to multiple deprivation cycles can partially overcome a genetic predisposition of NP, LAD-1, and LAD-2 rats for low alcohol consumption. In addition, the findings suggest that genetic control of low alcohol consumption in rats is not associated with the inability to display an ADE.

Elimination of Dietary Gluten and Development of Type 1 Diabetes in High Risk Subjects

Removal of dietary gluten is associated with a lower frequency of type 1 diabetes (T1D) in patients with celiac disease. Therefore, we performed a pilot study in which seven islet-antibody-positive first degree relatives of patients with T1D were placed on a gluten-free diet for 12 months, followed by gluten re-exposure for 12 months, to investigate whether this could reduce levels of circulating autoantibodies. We found that islet autoantibody levels at the end of the gluten-free period were not different to those before the commencement of the diet nor to antibody levels at the end of the gluten re-exposure period. In the present study, we have followed the 7 children formerly placed on a gluten-free diet for the manifestation of T1D for up to 5 years (mean follow-up time after fulfilling inclusion criteria: 4.8 years, SE 0.82 years) and compared them to 30 siblings and offspring of patients with T1D with similar characteristics to the intervention group (mean follow-up time: 5 years, SE 0.62 years). The cumulative 5-year risk of T1D in the intervention group did not differ from that in the prediabetic control group (42.9%, 95 CI (6.3-79.5%) vs. 49.7%, 95 CI (30.9-68.5%), p=0.87, log-rank test). These findings suggest that removing gluten from the diet over a period of one year is effective neither in the short nor in the long term in high-risk prediabetic individuals with a fully activated immune response to different islet antigens close to manifestation of T1D. These and recent data showing that exposure to dietary gluten in offspring of mothers and fathers with T1D very early in life is associated with an increased risk of developing islet antibodies also suggest that removal of dietary gluten should be tested as early as possible in children with an increased risk of islet autoimmunity, i.e. before an immune response to islet antigens is established.

Pavlovian conditioning of endotoxin-tolerance in rats

The most fascinating example of the bi-directional interaction between the central nervous system (CNS) and immune system is the behavioral conditioning of immune functions. We therefore investigated the behavioral conditioning of lipopolysaccharide (LPS)-induced endotoxin tolerance using the taste aversion paradigm. The conditioned stimulus (CS) saccharin was paired with the unconditioned stimulus (UCS) LPS over a five (CONDl) or four (COND2) days learning trial. Controls received drinking water with (SHAM) or without (UNT) LPS. Endotoxin tolerance was tested by determination of LPS-induced tumor necrosis factor (TNF)-α release. After the avoidance of the induced endotoxin-tolerance the CS saccharin was re-presented in all experimental groups. A the end of the re-exposure period a complete endotoxin tolerance was noticed in the CONDl- and COND2-group. In contrast, no effect of saccharin administration was observed in the SHAM- or UNT-group. Our data demonstrate for the first time the behavioral conditioning of endotoxin tolerance. Furthermore, these results contribute new aspects to the mechanisms underlying the development and modulation of endotoxin tolerance.

Increased incidence of dyskinesias and other behavioral effects of re-exposure to neuroleptic treatment in social colonies of Cebus apella monkeys.

Typical neuroleptic medications are still administered to as many as 40% of patients receiving antipsychotic treatment in the US. Intermittent administration or interruption of long-term neuroleptic medication for schizophrenia may increase the incidence of human tardive dyskinesias, and similarly may produce increasingly marked motor side-effects, parkinsonism, and other behavioral pathologies in non-human primates. Given these similarities, we addressed the issue of prolonged and intermittent typical neuroleptic treatment and dopaminergic function during a 5-year, multi-phase study with social colonies of Cebus apella monkeys. In the previously reported phase 1, we examined the effects of 48 weeks of exposure to, followed by withdrawal from, fluphenazine decanoate (FPZ). Phase 3 reported here examined the effects of 18 weeks of re-exposure to FPZ in these same monkeys, 91 weeks after discontinuation of their phase 1 FPZ treatment. Analysis of blood plasma FPZ indicated levels of 0.22+/-0.08 ng/ml for the six injections during the re-exposure period (n=54), comparable to the 0.24+/-0.07 ng/ml levels measured during our original treatment with this dose. Acute dyskinesias and dystonias increased by 300% upon re-exposure to FPZ; 15 of 18 FPZ-treated animals exhibited oral-buccal dyskinesias and all exhibited torticollis or retrocollis. Retreatment with FPZ was also associated with highly significant reductions in Self- and Environment-Directed Behavior and Directed Affiliation, effects similar to those seen during the original phase 1 FPZ treatment. Although FPZ re-treatment was associated with a significant reduction in Directed Aggression (an effect that was more robust than that observed during phase 1), in phase 3, we again observed an increase in Directed Aggression during early drug discontinuation when animals were in a stress-inducing situation. These results both support our phase 1 conclusion that typical neuroleptic medications may contribute to negative symptoms of schizophrenia and provide additional evidence for the possibility of increased aggression in stressful situations when medication is discontinued. Additionally, the results indicate that intermittent treatment with typical neuroleptics may dramatically increase the incidence of dystonias and dyskinesias.