Laura L. Murphy

Acute effects of δ-9-tetrahydrocannabinol on dopaminergic activity in several rat brain areas

Abstract In this work, we examined the acute effects of two doses of δ-9-tetrahydrocannabinol (THC) on several pre- and postsynaptic biochemical measures of dopaminergic activity in the striatum, limbic forebrain, and hypothalamic-anterior pituitary area of adult male rats. The exposure to a low dose of THC (0.5 mg/kg bw) decreased the number of striatal D 2 dopaminergic binding sites, but did not affect their affinity. Treatment with a higher dose of THC was ineffective. In addition, both doses decreased the number of D 1 dopaminergic binding sites in the limbic forebrain without changing their affinity. We did not find any changes in the dopamine (DA) or L -3,4-dihydroxyphenylacetic acid (DOPAC) content, or in the DOPAC/DA ratio, in either the striatum or limbic forebrain. THC treatment produced a dose-related decline in plasma prolactin (PRL) levels. Furthermore, both the basal and DA-inhibited in vitro release were PRL were reduced in animals exposed to THC in a dose-dependent manner. This inhibitory effect of THC on PRL release was accompanied by a decreased DOPAC/DA ratio in the medial basal hypothalamus that, in turn, may be a result of the fall in PRL levels rather than a direct action of the drug. These data show that acute exposure to THC can alter brain dopaminergic neurotransmission. Our results suggest that the reduction of PRL release following THC exposure, both in vivo and in vitro, might be elicited by a direct action of THC on the pituitary.

Effects of delta-9-tetrahydrocannabinol exposure on adrenal medullary function: evidence of an acute effect and development of tolerance in chronic treatments.

Previous studies have shown that the secretion of several stress-related hormones can be altered by exposure to marihuana or its purified constituents. The purpose of this study was to examine changes in adrenal medullary function caused by acute, subchronic and chronic treatments with two different doses of delta-9-tetrahydrocannabinol (THC). Acute exposure to THC caused a significant decrease in the adrenal medulla contents of both norepinephrine (NE) and epinephrine (E) and a significant increase in the E/NE ratio. These effects were mainly observed with the highest dose of THC, but they were not accompanied by a statistically significant decrease in adrenal medulla tyrosine hydroxylase activity, the rate-limiting enzyme in the catecholamine (CA) synthesis. These effects disappeared after seven or fourteen days of a daily THC treatment, which suggests the development of tolerance to this drug. Analysis of plasma PRL, ACTH and corticosterone levels showed some THC-related changes in these hormones. THC-induced modifications in ACTH and corticosterone were not in parallel to the changes in the adrenal medulla function, whereas those effects of acute THC on PRL release were statistically correlated with decreases of CA contents following acute THC. In conclusion, acute exposure to THC caused an alteration in the adrenal medullary function, reflected by a fall in endogenous stores of both CAs which could influence the adrenal medullary response to stress situations. This acute effect of THC could be mediated by the pituitary secretion of PRL, although the possibility of an effect directly exerted on the adrenal medulla chromaffin cells should be also considered.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of Δ9-THC on VIP-induced prolactin secretion in anterior pituitary cultures : evidence for the presence of functional cannabinoid CB1 receptors in pituitary cells

Peripheral administration of cannabinoid CB1 receptor agonists to laboratory rats induce a brief rise in plasma prolactin (PRL) levels followed by a prolonged decrease in PRL secretion from the pituitary. While the inhibitory component of this biphasic response depends on the cannabinoid-induced activation of dopamine release from hypothalamic terminals located in the median eminence, the neurobiological mechanisms underlying the activation phase of PRL release remains to be explained. In the present study the possible direct effect of the cannabinoid receptor agonist delta9-Tetrahydrocannabinol (THC) on prolactin secretion and cAMP accumulation was examined in anterior pituitary cultures. THC (0.1 and 1 microM) increased cAMP levels, and induced PRL release (1 and 10 mu). THC did not affect vasoactive intestinal peptide (VIP, 0.5 microM) induced cAMP accumulation in pituitary cultures, showing additive effects at THC 1 microM concentration. However, THC did prevent VIP-dependent increases in prolactin secretion. These results indicate that THC, through a direct pituitary action, activates both the synthesis of cAMP and PRL release and interferes with intracellular mechanisms involved in PRL secretion by VIP. These actions could be mediated through cannabinoid CB1 receptors which were found to be present in anterior pituitary cells, including lactotrophs, as revealed by immunocytochemistry with a specific polyclonal antibody raised against the CB1 receptor protein.

American Ginseng Inhibits Induced COX-2 and NFKB Activation in Breast Cancer Cells

BACKGROUND Epidemiologic evidence suggests reduced breast cancer mortality in users of American Ginseng (AG) (Panax quinquefolium). We hypothesized that AG extract decreases proliferation of human breast cancer cells via an anti-inflammatory effect applicable to the prevention of breast and other cancers. MATERIAL AND METHODS A defined lyophilized aqueous extract of AG (LEAG) was dissolved in DMSO 1mg/mL, and serially diluted in saline. The cell lines MDA MB 231 and MCF7 were stimulated with the phorbol ester PDBu and treated with 100-500 mcg/mL LEAG. Proliferation was measured by MDA assay. Induced COX-2 expression was assayed by ELISA. Activation of NFkappaB by phosphorylation of the p65 subunit was quantified by CASE (cellular activation of signaling ELISA). RESULTS Both cell lines had reduced proliferation when treated with LEAG. PDBu stimulation of MDA MB 231 increased expression of the COX-2 protein 20-fold at 48 hours (P<0.005). COX-2 protein expression remained at baseline concentrations in PDBu- treated MDA MB 231 cells exposed to 100 mcg/mL LEAG. The CASE assay showed a 4-fold increase in p65 activation 24 hours after PDBu treatment in normal medium, while phosphorylated p65 dropped below baseline in the cells treated with PDBu plus LEAG. CONCLUSION In MDA MB 231, COX-2 was inducible with PDBu. This induced COX-2 expression was blocked by 100 microgram/mL LEAG in a time course consistent with the decline in the activated p65 subunit of NFkappaB. These results provide an anti-inflammatory mechanism for a possible anti-cancer effect of American Ginseng.

Imaging the urokinase plasminongen activator receptor in preclinical breast cancer models of acquired drug resistance.

Subtype-targeted therapies can have a dramatic impact on improving the quality and quantity of life for women suffering from breast cancer. Despite an initial therapeutic response, cancer recurrence and acquired drug-resistance are commonplace. Non-invasive imaging probes that identify drug-resistant lesions are urgently needed to aid in the development of novel drugs and the effective utilization of established therapies for breast cancer. The protease receptor urokinase plasminogen activator receptor (uPAR) is a target that can be exploited for non-invasive imaging. The expression of uPAR has been associated with phenotypically aggressive breast cancer and acquired drug-resistance. Acquired drug-resistance was modeled in cell lines from two different breast cancer subtypes, the uPAR negative luminal A subtype and the uPAR positive triple negative subtype cell line MDA-MB-231. MCF-7 cells, cultured to be resistant to tamoxifen (MCF-7 TamR), were found to significantly over-express uPAR compared to the parental cell line. uPAR expression was maintained when resistance was modeled in triple-negative breast cancer by generating doxorubicin and paclitaxel resistant MDA-MB-231 cells (MDA-MB-231 DoxR and MDA-MB-231 TaxR). Using the antagonistic uPAR antibody 2G10, uPAR was imaged in vivo by near-infrared (NIR) optical imaging and 111In-single photon emission computed tomography (SPECT). Tumor uptake of the 111In-SPECT probe was high in the three drug-resistant xenografts (> 46 %ID/g) and minimal in uPAR negative xenografts at 72 hours post-injection. This preclinical study demonstrates that uPAR can be targeted for imaging breast cancer models of acquired resistance leading to potential clinical applications.

Curcumin binds tubulin, induces mitotic catastrophe, and impedes normal endothelial cell proliferation.

Curcumin, a component of turmeric spice that imparts flavor and color to curry, is thought to possess anti-inflammatory and antioxidant properties in biological tissues. However, while such efficacies have been described in the context of carcinogenesis, the impact of curcumin on normal cell cycle regulation is poorly understood. Here, we provide evidence of curcumin toxicity in proliferating bovine aortic endothelial cells, at concentrations relevant to the diet and below those previously reported in cancer models. Upon confirming curcumins ability to upregulate hemeoxygenase-1 in a dose-dependent fashion, we found the minimally efficacious curcumin concentration to also inhibit endothelial cell DNA synthesis. Moreover, curcumin concentrations below the minimum 2 μM threshold required to induce hemeoxygenase-1 bound tubulin protein in vitro and triggered hallmark evidence of mitotic catastrophe in vivo. Concentrations as low as 0.1 μM curcumin led to disproportionate DNA segregation, karyorrhexis, and micronucleation in proliferating endothelial cells. While suggesting a mechanism by which physiological curcumin concentrations inhibit cell cycle progression, these findings describe heretofore unappreciated curcumin toxicity with potential implications for endothelial growth, development, and tissue healing.

Phytonutrients Differentially Stimulate NAD(P)H:Quinone Oxidoreductase, Inhibit Proliferation, and Trigger Mitotic Catastrophe in Hepa1c1c7 Cells

UNLABELLED Phytonutrients have rapidly emerged as natural food chemicals possessing multifaceted biological actions that may support beneficial health outcomes. Among the vast array of phytonutrients currently being studied, sulforaphane, curcumin, quercetin, and resveratrol have been frequently reported to stimulate the expression of endogenous detoxification enzymes and may thereby facilitate the neutralization of otherwise harmful environmental agents. Some of these same phytonutrients, however, have also been implicated in disrupting normal cell proliferation and hence may possess toxic properties in and of themselves. In this study, we characterize the respective minimum threshold concentrations of the aforementioned phytonutrients in Hepa1c1c7 cells that stimulate NAD(P)H quinone oxidoreductase (NQO1), a key enzyme in the hepatic neutralization of menadione, other biological oxidants, and some environmental carcinogens. Moreover, our findings demonstrate that relatively low concentrations of either sulforaphane or curcumin significantly (P < .05) increase NQO1 protein expression and activity without triggering G2/M cell cycle arrest or mitotic catastrophe. The minimal quercetin concentration inducing NQO1, however, was 100-fold higher than that which disrupted mitosis. Also, while resveratrol modestly stimulated NQO1, the minimally effective resveratrol concentration concomitantly induced evidence of cellular apoptosis. Taken together, these findings indicate that only particular phytonutrients are likely efficacious in upregulating NQO1 activity without also leading to hepatic cytotoxicity.