Cannabinoids and the hippocampal glucocorticoid receptor: recent findings and possible significance.

It has long been recognized that cannabinoids, including delta 9-tetrahydrocannabinol (THC), the major psychoactive substance of marijuana, bear structural similarities to steroid hormones. The hippocampal region of the brain is particularly rich in glucocorticoid receptors (GCRs), and the region also displays dense autoradiographic binding by synthetic cannabinoids. The present report summarizes studies conducted on cannabinoid interaction with hippocampal GCRs, both in vivo and in vitro. Young rats treated for 8 months with THC displayed anatomic and cellular changes in the hippocampus similar to those seen in older, untreated rats, or in rats treated with high levels of glucocorticoids. Binding of [3H]dexamethasone in cytosol prepared from adrenalectomized rat hippocampus was reduced in the presence of 100-fold molar excess of unlabeled THC. However, further increases of THC concentration, to 20,000-fold excess, could displace no more than 50% of radiolabeled dexamethasone. Scatchard analysis of the binding produced a parallel competition plot for THC, versus the plot for dexamethasone, which may reflect a noncompetitive or allosteric interaction with hippocampal GCR. Cannabidiol, a nonpsychoactive cannabinoid, displayed less competition than THC in all parameters. Treatment of adrenalectomized rats for 14 days with 10 mg/kg THC produced down-regulation of hippocampal GCR binding in a manner also reported following high glucocorticoid administration. Although an initial oral administration of THC to intact rats stimulated release of plasma corticosterone, daily repetition of treatment for 7 and 14 days failed to elicit further corticosterone secretion. Taken together, the results indicate that THC may possess some agonist-like properties of glucocorticoids at the hippocampal GCR site.(ABSTRACT TRUNCATED AT 250 WORDS)     

Long-term effects of translocation and release numbers on fine-scale population structure among coho salmon (Oncorhynchus kisutch)

Management actions, such as translocations, reintroductions and supportive breeding, can have both negative and positive effects on population recovery. Several studies have examined the incidence of introgression following such actions, but few studies have explored the effect of release numbers on gene flow between closely related recipient populations. We examined population structure of coho salmon in Puget Sound (Washington State, USA) to evaluate the relationship between the number of individuals transferred between rivers, and the number released within rivers, on inter- and intrariver population divergence. Eleven microsatellite loci were surveyed in 23 hatchery and wild samples collected from 11 rivers within and one hatchery outside Puget Sound. Pairwise genetic divergences between most populations were significant, but the population structure could not be explained by an isolation-by-distance model (Mantel test, P > 0.05). In contrast, we detected significant hatchery influence on population structure. The numbers of fish transferred among rivers between 1952 and 2004 was negatively correlated with differentiation between rivers (partial Mantel test, P = 0.005) but not within rivers (t-test, P = 0.41). Number of fish released from hatcheries that collect broodstock locally was negatively correlated with population structure within rivers (t-test P = 0.002), and between nearby rivers (partial Mantel P = 0.04). Our results indicate that the population structure can, to some degree, be altered by the number of individuals transferred and by local release number of individuals in ongoing artificial propagation programs. The findings presented here emphasize the need to control the number of individuals that are either inadvertently introduced, or are deliberately released under conservation scenarios.     

Streptococcal serotype carbohydrate represents a novel class of type 2 antigen which is T-independent

Our studies reported here, fully characterize two unique type 2 antigens trinitrophenol (TNP)-M1 serotype carbohydrates (TNP-M1 g and TNP-M1 c) derived from streptococci, which fail to induce antibody responses in xid or neonatal mouse splenic cultures. These antigens generate brisk responses in normal spleen and Peyer's patch cell cultures of xid mice, all of which suggest that responses are elicited in the Lyb-3+, 5+ B subpopulation. The antibody responses to TNP-M1 g (and TNP-M1 c) are not dependent upon T cells. Furthermore, TNP-M1 carbohydrates induce anti-TNP plaque-forming (PFC) responses in cultures of small, resting splenic B cell populations without an added T cell requirement. Thus two categories of type 2 antigens are distinguished, one which requires T cells or derived factors, e.g., TNP-Ficoll, and a second TNP-carbohydrate antigen TNP-M1 that does not. Studies of the mitogenic and polyclonal B cell activation properties of M1 carbohydrates indicated that B cell proliferation is induced in both xid (Lyb-3-, 5-) and normal (Lyb-3-, 5- and Lyb-3+, 5+) splenic B cell subpopulations, but that differentiation to IgM synthesis fails to occur in the Lyb-3-, 5- B cell subpopulation. Thus M1 carbohydrates are unique probes that allow the selective induction of proliferation and differentiation of mature B cells that are presumably Lyb-3+, 5+. Because the M1 serotype carbohydrates induce polyclonal IgM synthesis and antigen-specific responses in only the mature B cell population in the absence of T cells, whereas TNP-Ficoll and other type 2 antigens require T cells or their derived factors, the Lyb-3+, 5+ B cell subpopulation may consist of a T cell-dependent and a T cell-independent compartment for responses to different carbohydrate type 2 antigens.     

Effects of melatonin on acetylsalicylic acid induced gastroduodenal and jejunal mucosal injury

We evaluated the effects of melatonin on acetylsalicylic acid (ASA) induced gastroduodenal and jejunal mucosal injury. We used 40 postpubertal rats divided randomly into five groups of eight animals. The control group consisted of untreated animals. The Mel group was injected intraperitoneally (i.p.) with 5 mg/kg melatonin. The ASA group was injected i.p. with 200 mg/kg ASA. The ASA + Mel group was injected i.p. with 5 mg/kg melatonin 45 min after administering 200 mg/kg ASA i.p. The Mel + ASA group was injected i.p. with 5 mg/kg melatonin 45 min before administering 200 mg/kg ASA i.p. We found no statistically significant differences in mean histopathological scores in the ASA + Mel group compared to the ASA group. ASA caused shortened villi and loss of the apical villus in the duodenum. The histopathological score was increased and villus height was decreased in the ASA group compared to untreated controls. Treatment with melatonin attenuated the histological damage. In the ASA group, occasional areas showed erosion of villi in the jejunum; however, differences in mean histopathological score in ASA group compared to the other groups were not statistically significant. Malondialdehyde (MDA), glutathione (GSH) and superoxide dismutase (SOD) activities were measured in stomach, duodenal and jejunum tissue. We found increased MDA activity in both stomach and duodenal tissues in the ASA group compared to the control group (p < 0.05). We found no statistically significant changes in MDA levels in jejunal tissue in the ASA group compared to the control group. We found no change in SOD activity in either stomach or duodenal tissues in the ASA group compared to the control group. We observed decreased SOD activity in jejunal tissue in the ASA group compared to the control group (p < 0.05). We detected no change in GSH activity in stomach, duodenal or jejunal tissues in the ASA group compared to the control group. The stomach damage was less in melatonin treated groups, but the lesions were not completely eliminated. The jejunum in the ASA group retained a nearly normal appearance. We found that melatonin exhibited some healing effects on ASA induced duodenal mucosal injury.     

Phylogenetic analysis supports horizontal transfer of P transposable elements

Nucleotide sequence comparisons were used to investigate the evolution of P transposable elements and the possibility that horizontal transfer has played a role in their occurrence in natural populations of Drosophila and other Diptera. The phylogeny of P elements was examined using published sequences from eight dipteran taxa and a new, partial sequence from Scaptomyza elmoi. The results from a number of different analyses are highly consistent and reveal a P-element phylogeny that contradicts the phylogeny of the species. At least three instances of horizontal transfer are necessary to explain this incongruence, but other explanations cannot be ruled out at this time.      

Limited Introgression between Rock-Wallabies with Extensive Chromosomal Rearrangements

Chromosome rearrangements can result in the rapid evolution of hybrid incompatibilities. Robertsonian fusions, particularly those with monobrachial homology, can drive reproductive isolation amongst recently diverged taxa. The recent radiation of rock-wallabies (genus Petrogale) is an important model to explore the role of Robertsonian fusions in speciation. Here, we pursue that goal using an extensive sampling of populations and genomes of Petrogale from north-eastern Australia. In contrast to previous assessments using mitochondrial DNA or nuclear microsatellite loci, genomic data are able to separate the most closely related species and to resolve their divergence histories. Both phylogenetic and population genetic analyses indicate introgression between two species that differ by a single Robertsonian fusion. Based on the available data, there is also evidence for introgression between two species which share complex chromosomal rearrangements. However, the remaining results show no consistent signature of introgression amongst species pairs and where evident, indicate generally low introgression overall. X-linked loci have elevated divergence compared with autosomal loci indicating a potential role for genic evolution to produce reproductive isolation in concert with chromosome change. Our results highlight the value of genome scale data in evaluating the role of Robertsonian fusions and structural variation in divergence, speciation, and patterns of molecular evolution.     

The pituitary-gonadal axis before puberty: Effects of various estrogenic steroids in the ovariectomized rat

A series of experiments were conducted to determine the effects of several estrogens upon FSH and LH secretion in immature ovariectomized rats. Groups of animals were castrated at 26 days of age and treated for 5 days post-operatively with various dosages of one of the following steroids: estrad1ol-17β(E₂), estradiol benzoate (EB), estrone (E₁), equilenin (EQ), 17α-ethinyl estradiol (EE), or mestranol (ME). Uterine weights were recorded and blood taken for radioimmunoassay.Estradiol was able to suppress both FSH and LH within a “physiologic dosage range” (PDR), wherein both gonadotropins were suppressed to intact control levels by a dose which did not stimulate uterine weight higher than intact control weight. EB and ME suppressed LH but not FSH at the PDR; the other steroids suppressed at higher than PDR doses. LH was preferentially suppressed, as compared to FSH, by all 6 steroids. By biological potency the order of activity was E₂ = EE ? EB ? ME ? E₁ ? EQ. For relative ability to suppress FSH (compared to LH), the order was E₁ or E₂, ? ME ?EB ? EE ? EQ. At higher doses (near maximum uterine stimulation), e₁, E₂ and EE produced higher FSH and LH than suppressed levels seen at lower doses; a pharmacologie dosage of E₂ caused re-suppression of both gonadotropins.Results indicate that a feedback system is present before puberty and this system is sensitive to very low levels of estrogens. Likewise, there is a potential for positive feedback present for higher estrogen levels, and a complete suppression occurs at pharmacologie levels. There appears to be a significant discrepancy between the biologic activity (by uterine weight) of estrogens and their ability to affect gonadotropin     

Increased hyaluronan fragmentation during pulmonary ischemia

Hyaluronan (HA), a glycosaminoglycan critical to the lung extracellular matrix, has been shown to dissociate into low-molecular-weight (LMW) HA fragments following exposure to injurious stimuli. In the present study we questioned whether lung HA changed during ischemia and whether changes had an effect on subsequent angiogenesis. After left pulmonary artery ligation (LPAL) in mice, we analyzed left lung homogenates immediately after the onset of ischemia (0 h) and intermittently for 14 days. The relative expression of HA synthase (HAS)1, HAS2, and HAS3 was determined by real-time RT-PCR, total HA in the lung was measured by an ELISA-like assay, gel electrophoresis was performed to determine changes in HA size distribution, and the activity of hyaluronidases was determined by zymography. A 50% increase in total HA was measured 16 h after the onset of ischemia and remained elevated for up to 7 days. Furthermore, a fourfold increase in LMW HA fragments (495-30 kDa) was observed by 4 h after LPAL. Both HAS1 and HAS2 showed increased expression 4-16 h after LPAL, yet no changes were seen in hyaluronidase activity. These results suggest that both HA fragmentation and activation of HA synthesis contribute to increased HA levels during lung ischemia. Delivery of LMW HA fragments in an in vitro tube formation assay or directly to the ischemic mouse lung in vivo both resulted in increased angiogenesis. We conclude that ischemic injury results in matrix fragmentation, which leads to stimulation of neovascularization.     

A Phase I Double Blind,Placebo-Controlled,Randomized Study of the Safety and Immunogenicity of Electroporated HIV DNA with or without Interleukin 12 in Prime-Boost Combinations with an Ad35 HIV Vaccine in Healthy HIV-Seronegative African Adults

Background

Strategies to enhance the immunogenicity of DNA vaccines in humans include i) co-administration of molecular adjuvants, ii) intramuscular administration followed by in vivo electroporation (IM/EP) and/or iii) boosting with a different vaccine. Combining these strategies provided protection of macaques challenged with SIV; this clinical trial was designed to mimic the vaccine regimen in the SIV study.

Methods

Seventy five healthy, HIV-seronegative adults were enrolled into a phase 1, randomized, double-blind, placebo-controlled trial. Multi-antigenic HIV (HIVMAG) plasmid DNA (pDNA) vaccine alone or co-administered with pDNA encoding human Interleukin 12 (IL-12) (GENEVAX IL-12) given by IM/EP using the TriGrid Delivery System was tested in different prime-boost regimens with recombinant Ad35 HIV vaccine given IM.

Results

All local reactions but one were mild or moderate. Systemic reactions and unsolicited adverse events including laboratory abnormalities did not differ between vaccine and placebo recipients. No serious adverse events (SAEs) were reported. T cell and antibody response rates after HIVMAG (x3) prime—Ad35 (x1) boost were independent of IL-12, while the magnitude of interferon gamma (IFN-γ) ELISPOT responses was highest after HIVMAG (x3) without IL-12. The quality and phenotype of T cell responses shown by intracellular cytokine staining (ICS) were similar between groups. Inhibition of HIV replication by autologous T cells was demonstrated after HIVMAG (x3) prime and was boosted after Ad35. HIV specific antibodies were detected only after Ad35 boost, although there was a priming effect with 3 doses of HIVMAG with or without IL-12. No anti-IL-12 antibodies were detected.

Conclusion

The vaccines were safe, well tolerated and moderately immunogenic. Repeated administration IM/EP was well accepted. An adjuvant effect of co-administered plasmid IL-12 was not detected.

Trial Registration

ClinicalTrials.gov NCT01496989