Germinal epimutation of Fragile Histidine Triad (FHIT) gene is associated with progression to acute and chronic adult T-cell leukemia diseases.

BACKGROUND: Human T cell Leukemia virus type 1 (HTLV-I) is etiologically linked to adult T cell leukemia/lymphoma (ATL) and an inflammatory neurodegenerative disease called HTLV-I-associated myelopathy or tropical spastic paraparesis (HAM/TSP). The exact genetic or epigenetic events and/or environmental factors that influence the development of ATL, or HAM/TSP diseases are largely unknown. The tumor suppressor gene, Fragile Histidine Triad Diadenosine Triphosphatase (FHIT), is frequently lost in cancer through epigenetic modifications and/or deletion. FHIT is a tumor suppressor acting as genome caretaker by regulating cellular DNA repair. Indeed, FHIT loss leads to replicative stress and accumulation of double DNA strand breaks. Therefore, loss of FHIT expression plays a key role in cellular transformation. METHODS: Here, we studied over 400 samples from HTLV-I-infected individuals with ATL, TSP/HAM, or asymptomatic carriers (AC) for FHIT loss and expression. We examined the epigenetic status of FHIT through methylation specific PCR and bisulfite sequencing; and correlated these results to FHIT expression in patient samples. RESULTS: We found that epigenetic alteration of FHIT is specifically found in chronic and acute ATL but is absent in asymptomatic HTLV-I carriers and TSP/HAM patients' samples. Furthermore, the extent of FHIT methylation in ATL patients was quantitatively comparable in virus-infected and virus non-infected cells. We also found that longitudinal HTLV-I carriers that progressed to smoldering ATL and descendants of ATL patients harbor FHIT methylation. CONCLUSIONS: These results suggest that germinal epigenetic mutation of FHIT represents a preexisting mark predisposing to the development of ATL diseases. These findings have important clinical implications as patients with acute ATL are rarely cured. Our study suggests an alternative strategy to the current "wait and see approach" in that early screening of HTLV-I-infected individuals for germinal epimutation of FHIT and early treatment may offer significant clinical benefits.

IL-6 deficiency attenuates p53 protein accumulation in aged male mouse hippocampus.

Our earlier studies demonstrated slower age-related memory decline in IL-6-deficient than in control mice. Therefore, in the present study we evaluated the effect of IL-6 deficiency and aging on expression of p53, connected with accumulation of age-related cellular damages, in hippocampus of 4- and 24-month-old IL-6-deficient C57BL/6J (IL-6KO) and wild type control (WT) mice. The accumulation of p53 protein in hippocampus of aged IL-6KO mice was significantly lower than in aged WT ones, while p53 mRNA level was significantly higher in IL-6-deficient mice, what indicates that the effect was independent on p53 transcription. Presence of few apoptotic cells in hippocampal dentate gyrus and lack of changes in levels of pro-apoptotic Bax, antiapoptotic Bcl-2, as well as in p21 protein in aged animals of both genotypes, points to low transcriptional activity of p53, especially in aged WT mice. Because the amount of p53 protein did not correlate with the level of Mdm2 protein, its main negative regulator, other than Mdm2-dependent mechanism was involved in p53 build-up. Significantly higher mRNA levels of autophagy-associated genes: Pten, Tsc2, and Dram1 in IL-6KO mice, in conjunction with significantly lower amount of Bcl-2 protein in 4-month-old IL-6KO mice, suggests that lack of IL-6/STAT3/Bcl-2 signaling could account for better autophagy performance in these mice, preventing excessive accumulation of proteins. Taken together, attenuated p53 protein build-up, absence of enhanced apoptosis, and transcriptional up-regulation of autophagy-associated genes imply that IL-6 deficiency may protect hippocampus from age-related accumulation of cellular damages.

Significance of IL-6 Deficiency in Recognition Memory in Young Adult and Aged Mice.

Chronic peripheral elevation of interleukin 6 (IL-6) in humans is associated with cognitive deficits. 4- and 24-month-old IL-6-deficient C57BL/6J (IL-6KO) and reference wild-type (WT) mice were tested in an object recognition test. Discrimination ratios and recognition indexes were significantly lower in 4-month-old IL-6KO and in 24-month-old WT mice vs 4-month-old WT animals. Their discrimination ratios had negative values and recognition indexes were below 50% indicating inability to differentiate the novel from the familiar object after 1-hour delay. In 24-month-old IL-6KO mice recognition index reached 53.17% indicating that their recognition memory was not worsened with age in comparison with younger IL-6-deficient animals. Results of holeboard and elevated plus maze indicated that this effect was memory specific. Inborn IL-6 deficiency attenuated recognition memory in 4-month-old mice and did not altered recognition memory in aged animals. IL-6 signalling may constitute a target for development of the protection against memory disturbances connected with IL-6 overexpression.

IL-6 deficiency alters spatial memory in 4- and 24-month-old mice.

Significance of interleukin 6 (IL-6) deficiency in cognitive processes was evaluated in 4- and 24-month-old C57BL/6J IL-6-deficient (IL-6 KO) and control (WT) mice in Morris water maze (MWM), holeboard test (HB) and elevated plus maze (EPM). During 3-day learning escape latency time (ELT) was longer in IL-6 KO than in WT mice, however their swimming was slower, floating longer, and path length did not differ. The comparison of ELT and the distance traveled between the first and the third learning day within each group revealed significant decrease of ELT in all groups with the highest difference in 4-month-old WT mice, and significant decrease of distance traveled only in both groups of WT mice. In a single probe trial, performed 24 h after the last learning session, there were no major differences in the absolute values of ELT, but ELT turned out to be significantly shorter in both IL-6 KO groups, when it was compared to the ELT on the last learning day, indicating on better memory retrieval. In HB test only significant increase in number of rearings in aged WT mice, and in EPM significant prolongation of open arm time and higher number of open arm entries in 4-month-old IL-6 KO mice were observed. Results of HB and EPM tests showed that alterations of learning and reference memory observed in MWM were specific to cognition. Attenuation of learning ability in young adult IL-6-deficient mice assessed in MWM suggests that physiological level of IL-6 is involved in mechanisms engaged in proper memory formation, and it may also indicate on the importance of IL-6 signaling in brain development. Maintained on similar level in both 4- and 24-month-old IL-6 KO mice learning ability and its attenuation in 24-month-old vs 4-month-old WT mice indicates on slower age-related memory decline in mice not expressing IL-6. Better performance of IL-6 KO mice in the probe trial points to their reference memory improvement and may also indicate that IL-6 plays a role in mechanism responsible for cognitive flexibility.

Facilitatory Effect of IL-6 Deficiency on Long-Term Spatial Memory in Young Adult Mice.

The significance of interleukin 6 (IL-6) in long-term reference memory was tested in the Morris water maze (MWM) in 4-month-old C57BL/6J IL-6-deficient (IL-6 KO) and control mice. Three-day learning measured by escape latency time to find the hidden platform was comparable in both genotypes. In a single probe trial performed 7 days later, without the platform, latency to the platform site and path length to the target place were significantly shorter (p < 0.05 and p < 0.02, respectively), and platform-site crossovers more frequent (p < 0.05) in IL-6 KO mice. The swimming speed in IL-6 KO mice was significantly lower during learning (p = 0.0025) but not in the probe trial. Lack of differences between genotypes in a hole-board and in an elevated plus maze indicates that the observed effects were memory specific. The facilitatory effect of IL-6 deficiency on long-term reference memory in MWM indicates that IL-6 plays a role in consolidation process.

Co-dependence of HTLV-1 p12 and p8 functions in virus persistence.

HTLV-1 orf-I is linked to immune evasion, viral replication and persistence. Examining the orf-I sequence of 160 HTLV-1-infected individuals; we found polymorphism of orf-I that alters the relative amounts of p12 and its cleavage product p8. Three groups were identified on the basis of p12 and p8 expression: predominantly p12, predominantly p8 and balanced expression of p12 and p8. We found a significant association between balanced expression of p12 and p8 with high viral DNA loads, a correlate of disease development. To determine the individual roles of p12 and p8 in viral persistence, we constructed infectious molecular clones expressing p12 and p8 (D26), predominantly p12 (G29S) or predominantly p8 (N26). As we previously showed, cells expressing N26 had a higher level of virus transmission in vitro. However, when inoculated into Rhesus macaques, cells producing N26 virus caused only a partial seroconversion in 3 of 4 animals and only 1 of those animals was HTLV-1 DNA positive by PCR. None of the animals exposed to G29S virus seroconverted or had detectable viral DNA. In contrast, 3 of 4 animals exposed to D26 virus seroconverted and were HTLV-1 positive by PCR. In vitro studies in THP-1 cells suggested that expression of p8 was sufficient for productive infection of monocytes. Since orf-I plays a role in T-cell activation and recognition; we compared the CTL response elicited by CD4+ T-cells infected with the different HTLV-1 clones. Although supernatant p19 levels and viral DNA loads for all four infected lines were similar, a significant difference in Tax-specific HLA.A2-restricted killing was observed. Cells infected with Orf-I-knockout virus (12KO), G29S or N26 were killed by CTLs, whereas cells infected with D26 virus were resistant to CTL killing. These results indicate that efficient viral persistence and spread require the combined functions of p12 and p8.

Humoral immune response to HTLV-1 basic leucine zipper factor (HBZ) in HTLV-1-infected individuals.

BACKGROUND: Human T cell lymphotropic virus type 1 (HTLV-1) infection can lead to development of adult T cell leukemia/lymphoma (ATL) or HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) in a subset of infected subjects. HTLV-1 basic leucine zipper factor (HBZ) gene has a critical role in HTLV-1 infectivity and the development of ATL and HAM/TSP. However, little is known about the immune response against HBZ in HTLV-1-infected individuals. In this study, we examined antibody responses against HBZ in serum/plasma samples from 436 subjects including HTLV-1 seronegative donors, asymptomatic carriers (AC), ATL, and HAM/TSP patients using the luciferase immunoprecipitation system. RESULTS: Immunoreactivity against HBZ was detected in subsets of all HTLV-1-infected individuals but the test did not discriminate between AC, ATL and HAM/TSP. However, the frequency of detection of HBZ-specific antibodies in the serum of ATL patients with the chronic subtype was higher than in ATL patients with the lymphomatous subtype. Antibody responses against HBZ were also detected in cerebrospinal fluid of HAM/TSP patients with anti-HBZ in serum. Antibody responses against HBZ did not correlate with proviral load and HBZ mRNA expression in HAM/TSP patients, but the presence of an HBZ-specific response was associated with reduced CD4+ T cell activation in HAM/TSP patients. Moreover, HBZ-specific antibody inhibited lymphoproliferation in the PBMC of HAM/TSP patients. CONCLUSIONS: This is the first report demonstrating humoral immune response against HBZ associated with HTLV-I infection. Thus, a humoral immune response against HBZ might play a role in HTLV-1 infection.

Requirement of the human T-cell leukemia virus p12 and p30 products for infectivity of human dendritic cells and macaques but not rabbits.

The identification of the genes necessary for human T-cell leukemia virus (HTLV-1) persistence in humans may provide targets for therapeutic approaches. We demonstrate that ablation of the HTLV-1 genes encoding p12, p30, or the HBZ protein, does not affect viral infectivity in rabbits and in this species, only the absence of HBZ is associated with a consistent reduction in virus levels. We observed reversion of the HTLV-1 mutants to the HTLV-1 wild-type genotype in none of the inoculated rabbits. In contrast, in macaques, the absence of HBZ was associated with reversion of the mutant virus to the wild-type genotype in 3 of the 4 animals within weeks from infection. Similarly, reversion to the wild type was observed in 2 of the 4 macaque inoculated with the p30 mutant. The 4 macaques exposed to the p12 knock remained seronegative, and only 2 animals were positive at a single time point for viral DNA in tissues. Interestingly, we found that the p12 and the p30 mutants were also severely impaired in their ability to replicate in human dendritic cells. These data suggest that infection of dendritic cells may be required for the establishment and maintenance of HTLV-1 infection in primate species.

In vivo genetic mutations define predominant functions of the human T-cell leukemia/lymphoma virus p12I protein.

The human T-cell leukemia/lymphoma virus type 1 (HTLV-1) ORF-I encodes a 99-amino acid hydrophobic membrane protein, p12(I), that affects receptors in different cellular compartments. We report here that proteolytic cleavage dictates different cellular localization and functions of p12(I). The removal of a noncanonical endoplasmic reticulum (ER) retention/retrieval signal within the amino terminus of p12(I) is necessary for trafficking to the Golgi apparatus and generation of a completely cleaved 8-kDa protein. The 8-kDa protein in turn traffics to the cell surface, is recruited to the immunologic synapse following T-cell receptor (TCR) ligation, and down-regulates TCR proximal signaling. The uncleaved 12-kDa form of p12(I) resides in the ER and interacts with the beta and gamma(c) chains of the interleukin-2 receptor (IL-2R), the heavy chain of the major histocompatibility complex (MHC) class I, as well as calreticulin and calnexin. Genetic analysis of ORF-I from ex vivo samples of HTLV-1-infected patients reveals predominant amino acid substitutions within ORF-I that affect proteolytic cleavage, suggesting that ER-associated functions of p12(I) may contribute to the survival and proliferation of the infected T cells in the host.

Interleukin 6 Knockout Inhibits Aging-Related Accumulation of p53 in the Mouse Myocardium.

Interleukin 6 (IL6) and p53 are linked by mutual regulatory mechanisms and are both upregulated in aging. The aim of this study was to evaluate the effects of aging and IL6 on expression of p53 in the mouse heart. Male C57BL6/J wild-type and IL6 knockout mice at the age of 4-5 months (young adult) and 24-30 months (old) were used. Myocardial expression of proteins such as p53, p21, Mdm2, and phospho-Akt/Akt was estimated using Western blotting and expression of p53 and p21 mRNA using real-time polymerase chain reaction. Expression of p53 protein was lower in IL6 knockout hearts than in wild-type hearts. Aging caused significant upregulation of p53 protein level; however, it was significantly higher in old wild-type hearts than in old IL6 knockout hearts (p < .05). Similar p53 mRNA levels in all groups implied IL6 influence on age-related proteasomal degradation of p53. Localization of p53 mainly in the extranuclear compartment and lack of p21 upregulation in aged hearts may suggest quenched transcriptional activity of p53 despite increased abundance of p53. We conclude that lack of IL6 attenuates expression of p53 protein in the hearts of young mice and diminishes its accumulation with aging by post-transcriptional mechanisms; however, this is not related to altered phenotype of aging heart.

Expression of collagens type I and V in healing rat's tympanic membrane.

OBJECTIVES: Our preliminary study performed on perforated rat's tympanic membrane (TM) using Rat Wound Healing RT2 Profiler PCR Array showed significantly increased levels of mRNA for collagens type I and V. Enhanced expression of those genes does not assure that their protein products are indeed present, and in what quantity. Therefore, this study was undertaken to analyze the collagen type I and V content in the healing TM. METHODS: Sixty rats were used, of which 10 served as controls and the others had their TM perforated. The experimental animals were divided into five subgroups on the basis of time points (03, 06, 09, 14, 20 day after injury). Videootoscopy and histology were employed to assess the morphology of the healing process. The expression of collagen type I and V was evaluated using Western blot analysis. Tissue localization of collagens was determined by the immunofluorescence method. RESULTS: The collagen type I expression was three times higher on the third day after injury and remained on that level for whole period of observation, up to day 20. The increase of the collagen type V expression was gradual, reaching the highest level on day 14 following injury. In comparison to the control TM statistically significant increase in the level of expression was observed starting from day 09 to the end of observation period. In healing TM immunofluorescent labeling of collagen type I and V was seen on the surface of remnants of previous lamina propria and in the loose proliferating fibrous tissue. On day 20 immunofluorescence was present mainly on the surface of thin connective tissue layers forming the scar in the place of previous perforation. CONCLUSION: Although the collagens type I and V are present only in subepithelial layer in the normal rat's TM they play significant role in TM healing process.

Impairment of recognition memory in interleukin-6 knock-out mice.

We have used transgenic male mice not expressing interleukin-6 (IL-6) [C57BL/6J(IL-6/-tm Kopf)] in object recognition test to assess the role of endogenous IL-6 in recognition memory. Wild-type controls showed better memory than IL-6 knock-out mice. Results of our experiment suggest that endogenous IL-6 may play an important role in the process of recognition memory in mice.

Facilitatory effect of AM281 on recognition memory in rats.

BACKGROUND: Our approach was to determine the influence of a single systemic administration of AM281, synthetic cannabinoid structurally similar to SR141716A, on recognition memory in rats. METHODS: To assess the influence of AM281 on acquisition of information the compound was given intraperitoneally once, at the doses of 0.1, 0.5, 1.0 or 2.0mg/kg, 15min before learning trial (T1) and in order to evaluate its influence on consolidation process AM281 was given at indicated doses, immediately after T1 trial in an object recognition test. Since cannabinoids may alter motor function and affect anxiety, the influence of AM281 on psychomotor activity and anxiety was evaluated in an open-field and elevated plus maze test, respectively. RESULTS: Administration of AM281 at the doses: 0.1, 0.5 and 1.0mg/kg significantly improved acquisition of information, while 0.1 and 0.5mg/kg of AM281 significantly facilitated consolidation process. Not only did AM281 not affect locomotor and exploratory activity, but also anxiety. CONCLUSION: This is the first evidence that AM281 exerts facilitatory effect on recognition memory in rats. This effect seems to be memory specific because no alterations in animals' psychomotor activity and anxiety were observed.

Analyses of HTLV-1 sequences suggest interaction between ORF-I mutations and HAM/TSP outcome.

The region known as pX in the 3' end of the human T-cell lymphotropic virus type 1 (HTLV-1) genome contains four overlapping open reading frames (ORF) that encode regulatory proteins. HTLV-1 ORF-I produces the protein p12 and its cleavage product p8. The functions of these proteins have been linked to immune evasion and viral infectivity and persistence. It is known that the HTLV-1 infection does not necessarily imply the development of pathological processes and here we evaluated whether natural mutations in HTLV-1 ORF-I can influence the proviral load and clinical manifestation of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). For that, we performed molecular characterization, datamining and phylogenetic analysis with HTLV-1 ORF-I sequences from 156 patients with negative or positive diagnosis for HAM/TSP. Our analyses demonstrated that some mutations may be associated with the outcome of HAM/TSP (C39R, L40F, P45L, S69G and R88K) or with proviral load (P34L and F61L). We further examined the presence of mutations in motifs of HBZ and observed that P45L mutation is located within the HBZ nuclear localization signal and was found more frequently between patients with HAM/TSP and high proviral load. These results indicate that some natural mutations are located in functional domains of ORF-I and suggests a potential association between these mutations and the proviral loads and development of HAM/TSP. Therefore it is necessary to conduct functional studies aimed at evaluating the impact of these mutations on the virus persistence and immune evasion.

Enhanced expression of hepatocyte growth factor in the healing of experimental acute tympanic membrane perforation.

OBJECTIVES: The present study was performed to investigate the expression of hepatocyte (HGF), epidermal (EGF) and vascular endothelial (VEGF) growth factors in the course of healing of experimental tympanic membrane (TM) perforations in rats. The goal was to explain the role of these growth factors in the healing process of TM and to assess the possibility of their future application as healing promoters. METHODS: Seventy rats were used, of which 10 served as controls and the others had their TM perforated. The experimental animals were divided into six subgroups on the basis of time points (01, 03, 05, 07, 09, 15 day after injury). Videootoscopy and histology were employed to assess the morphology of the healing process. The expressions of HGF, EGF and VEGF were evaluated using Western blot analysis. Tissue localization of HGF was determined by the immunofluorescence method. RESULTS: HGF was hardly detectable in normal TM; however, a significant increase was noted in its expression starting from the third day after injury throughout the follow-up period, with the highest level on day 05. The analysis of HGF tissue localization with immunofluorescence revealed diffuse staining in the cytoplasm of proliferating epithelial cells. The expression of EGF was elevated on the first day after injury, not reaching statistical significance, and then returned to the level observed in the control TM. No significant differences were noted in the expression of VEGF. CONCLUSION: High expression of HGF during the healing process of acute TM perforations makes it a promising candidate for further studies oriented towards its possible use in augmentation of TM healing.

Transcriptional and post-transcriptional regulation of CCN genes in failing heart.

BACKGROUND: CCN family of proteins has been implicated in various processes in cardiovascular physiology and pathology, including angiogenesis, regeneration and fibrosis. In this study we assessed long term changes of CCN1 and CCN2 gene products abundance in the failing ventricular myocardium. METHODS: Male, 12-14-weeks-old C57BL6/J and C57BL6/J (IL-6-/-) mice were used. To assess short term changes, a transient reversible ischemia model was utilized. Heart failure was caused by ligation of anterior descending coronary artery. The presence of systolic dysfunction was confirmed by echocardiography and left ventricular ANP RNA expression. Molecular analysis was performed on left ventricular samples from animals sacrificed 12-14 weeks after infarction. Western blotting and QT-PCR were used to investigate abundance of CCN proteins and RNAs, respectively. RESULTS: Short ischemia resulted in marked increase of CCN1 transcript. However, three months after myocardial infarction (MI), remote myocardium showed a markedly increased expression of CCN1 protein, but not RNA. In the case of CCN2, the RNA was distinctly up-regulated, whereas the protein presented only modest, non-significant increase in failing myocardium. Expression of CCN2 RNA closely correlated with expression of ANP. Long-term telmisartan administration after infarction decreased the expression of CCN1 protein. Interleukin 6 (IL-6) deficiency caused increased CCN2 protein abundance in control animals, but the difference was absent after MI. Infarction did not increase CCN1 protein in the hearts of IL-6 deficient mice. CONCLUSION: CCN genes are activated in heart failure. Their regulation is multidimensional both transcriptional and posttranscriptional. The involved pathways include angiotensin II and IL-6.

Enhancement of photosynthetic O2 evolution in Chlorella vulgaris under high light and increased CO2 concentration as a sign of acclimation to phosphate deficiency.

The photosynthetic oxygen evolution of Chlorella vulgaris (Beijer.) cells taken from phosphate-deficient (-P) and control cultures was measured during 8 days of culture growth. Under inorganic carbon concentration (50 microM) in the measuring cell suspension and irradiance (150 micromol m(-2) s(-1)), the same as during culture growth, there were no marked differences in the photosynthetic O2 evolution rate between the -P cells and the controls. The much slower growth of -P cultures indicated that the utilization of absorbed photosynthetically active radiation (PAR) in the CO2 assimilation and biomass production were in -P cells less efficient than in the controls. Alga cells under the phosphorus stress utilized more of the absorbed PAR in the nitrate reduction than the control cells. However, under conditions of more efficient CO2 supply (inorganic carbon concentration 150 microM, introducing of exogenous carbonic anhydrase to the measuring cell suspension) and under increased irradiance (500 micromol m(-2) s(-1)), the photosynthetic O2 evolution in -P cells reached a higher rate than in the controls. The results suggest that in -P cells the restricted CO2 availability limits the total photosynthetic process. But under conditions more favorable for the CO2 uptake and under high irradiance, the -P cells may reveal a higher photosynthetic oxygen evolution rate than the controls. It is concluded that an increased potential activity of the photosynthetic light energy absorption and conversion in the C. vulgaris cells from -P cultures is a sign of acclimation to phosphorus stress by a sun-type like adaptation response of the photosynthetic apparatus.

Preliminary immunohistochemical investigations of thyroid C cells in an experimental model of hyperthyroidism.

The role of the parafollicular (C) cells, the second most important cells in the thyroid gland, has not hitherto been clarified. They are considered to be disperse neuroendocrine cells of the APUD system and synthesise and release many of the regulatory peptides. Few publications are concerned with the evaluation of the structure and function of C cells in the thyroid gland or the probable relationship between these cells and the follicular cells in physiological and pathological conditions. For this reason immunohistochemical investigations were carried out into the activity of the C cells in rats in an experimental model of hyperthyroidism caused by chronic thyroxine influence. This C-cell activity was then evaluated. Differences in the quantity, distribution and calcitonin immunoreactivity of C cells were observed in hyperthyroid rats in comparison to the control group, together with a significant diminution of plasma TSH and calcitonin levels. Our preliminary study may indicate a functional interaction between follicular and parafollicular cells in the thyroid gland.

An immunohistochemical study of the thyroid parafollicular (C) cells in rats treated with cannabinoids - preliminary investigations.

The purpose of the present study was to evaluate the effect of a single intraperitoneal injection of a stable analogue of endogenous cannabinoid anandamide - R-(+)-methanandamide (2.5 mg/kg) and CP 55,940 (0.25 mg/kg), an egzogenous CB1 receptor-agonist, on the calcitonin (CT) immunoreactivity of the thyroid parafollicular (C) cells. Four hours after injection with both cannabinoids CT immunoreactivity, evaluated with an avidin-biotin peroxidase complex method by means of rabbit antibodies against CT, was seen to be enhanced in the parafollicular cells in comparison to those of the control group. In thyroids taken from cannabinoid-treated rats the majority of follicles, particularly those located peripherally were large in size, and had low epithelium. Moreover, dilatation of the blood vessels was observed. These changes were accompanied by a significant decrease in CT plasma level, without changes in calcium concentrations. This is the first evidence that a single injection of the cannabinoids R-(+)-methanandamide and CP 55,940 significantly decreases the activity of thyroid C cells.

CP55,940 attenuates spatial memory retrieval in mice.

BACKGROUND: Cannabinoids constitute a varied group of lipophilic substances able to infiltrate the blood-brain barrier and influence neuronal processes. Clinical observations supported by experimental data have revealed that these compounds exert a deleterious effect on cognitive processes. The present study was carried out to determine the influence of a single systemic administration of CP55,940, a potent synthetic agonist of cannabinoid receptors, on spatial memory retrieval assessed in a Morris water maze. METHODS: C57BL/6J male mice were submitted to three consecutive days of training to find a hidden platform in the water maze. CP55,940 was given intraperitoneally once, at doses of 0.025, 0.125 or 0.25mg/kg on the fourth day, 30min before testing memory retrieval, and in separate groups before testing psychomotor activity and anxiety level in a hole-board test. RESULTS: CP55,940 only at the highest dose of 0.25mg/kg significantly altered all parameters used to assess spatial memory. It increased the latency in the first crossing of the former platform location (target area), decreased the number of target area crossings and shortened the time spent in the target quadrant. Moreover, CP55,940 at doses of 0.25 and 0.125mg/kg attenuated motor and exploratory activity in hole-board test. CONCLUSION: Since the attenuated psychomotor activity after a dose of 0.125mg/kg did not interfere with memory retrieval, we assume that the impairment of spatial memory observed after the highest dose of CP55,940 (0.25mg/kg) was exerted by its influence on cognitive processes, however, the impact on locomotion could not be excluded.