An investigation of the genotoxic potential of a well-characterized yerba mate extract.

Yerba Mate (Ilex paraguariensis) is historically used as a beverage and its extracts are considered traditional medicine in South America. Extract use has been expanding to North American and European markets and the currently available genetic toxicology literature indicate discrepancies in genotoxicity findings for yerba mate. As botanical extract use expands, assumption in safety should be made with caution assuring a good understanding of the test material characterization. Authoritative agencies suggest a two-step paradigm to investigate genotoxicity, and this was implemented to evaluate the safety of yerba mate hydroxycinnamic acid extract. Four OECD compliant assays were employed: bacterial reverse mutation, in vitro micronucleus and a parallel in vivo micronucleus, and comet assay. No evidence of mutagenicity was observed in the in vitro Ames assay, but the results of an in vitro micronucleus study were inconclusive. However, oral gavage treatment of rats for the in vivo micronucleus and comet assays demonstrated negative findings. The results from this battery of tests, supports that this yerba mate hydroxycinnamic acid extract is not anticipated to pose genotoxicity concerns. A high-level comparison of results to other available genotoxicity literature on yerba mate is presented with emphasis on the importance of identity when drawing conclusions on botanicals.

A novel phenylaminotetralin (PAT) recognizes histamine H1 receptors and stimulates dopamine synthesis in vivo in rat brain.

A series of novel phenylaminotetralins (PATs) previously was shown to recognize discrete binding sites that are stereoselectively labeled by [3H]-(-)-trans-1-phenyl-3-N,N-dimethylamino-1,2,3,4-tetrahydronaphthalen e (H2-PAT) and highly localized in catecholaminergic nerve terminal regions in guinea pig forebrain. Furthermore, certain PATs stimulate tyrosine hydroxylase and dopamine synthesis in guinea pig and rat brain in vitro. In the current studies, we characterized sites labeled by [3H]-(-)-trans-H2-PAT and measured effects of PATs on dopamine synthesis in vivo in rat brain. [3H]-(-)-Trans-H2-PAT binds saturably (Bmax approximately 13 fmol/mg protein) and with high affinity (K(D) approximately 0.5 nM) to a single population of sites in rat brain. The ligand binding profile of [3H]-(-)-trans-H2-PAT labeled sites is very similar to histamine H1 receptors labeled with [3H]-mepyramine. After i.c.v. injection to rats, (+/-)-trans H2-PAT (4-40 nmoles/kg) stimulates dopamine synthesis (to about 180% of control levels) selectively in the limbic brain region nucleus accumbens vs. the extrapyramidal region striatum; this effect is fully blocked by (+/-)-cis-H2-PAT and the H1 antagonist triprolidine. At higher doses (> 40 nmoles/kg), the observed stimulation of dopamine synthesis is attenuated to control levels, likely due to activation of feedback mechanisms resulting from non-receptor mediated displacement of intraneuronal dopamine. We propose that PATs represent a novel class of ligands for H1 receptors that can modulate tyrosine hydroxylase activity and dopamine synthesis in the limbic region of mammalian forebrain.

Repeated exposure of adult rats to Aroclor 1254 causes brain region-specific changes in intracellular Ca2+ buffering and protein kinase C activity in the absence of changes in tyrosine hydroxylase.

Polychlorinated biphenyls (PCBs) are ubiquitous environmental contaminants, some of which may be neurotoxic. In vitro studies from this laboratory indicated that noncoplanar PCBs perturbed intracellular signal transduction mechanisms including Ca2+ homeostasis, receptor-mediated inositol phosphate production, and translocation of protein kinase C (PKC). In the present study, we examined the effects of PCBs in vivo by dosing adult male Long-Evans rats orally with Aroclor 1254 (0, 10, or 30 mg/kg/day; 5 days/week for 4 weeks) in corn oil. At 24 h after the last dose, rats were tested for motor activity in a photocell device for 30 min. Immediately, the rats were euthanized, blood was collected for thyroid hormone analysis, and brains were removed, dissected into regions (cerebellum, frontal cortex, and striatum), and subcellular fractions were obtained for neurochemical analysis. Following Aroclor 1254 treatment, body weight gain in the high-dose group was significantly lower than the control and low-dose groups. Horizontal motor activity was significantly lower in rats dosed with 30 mg/kg Aroclor 1254. Ca2+ buffering by microsomes was significantly lower in all three brain regions from the 30 mg/kg group. In the same dose group, mitochondrial Ca2+ buffering was affected in cerebellum but not in cortex or striatum. Similarly, total cerebellar PKC activity was decreased significantly while membrane-bound PKC activity was significantly elevated at 10 and 30 mg/kg. PKC activity was not altered either in cortex or the striatum. Neurotransmitter levels in striatum or cortex were slightly altered in PCB-exposed rats compared to controls. Furthermore, repeated oral administration of Aroclor 1254 to rats did not significantly alter forebrain tyrosine hydroxylase immunoreactivity or enzymatic activity. Circulating T4 (total and free) concentrations were severely depressed at both doses in Aroclor 1254-exposed rats compared to control rats, suggesting a severe hypothyroid state. These results indicate that (1) in vivo exposure to a PCB mixture can produce changes in second messenger systems that are similar to those observed after in vitro exposure of neuronal cell cultures; (2) second messenger systems seem to be more sensitive than alterations in neurotransmitter levels or tyrosine hydroxylase involved in dopamine synthesis during repeated exposure to PCBs; and (3) the observed motor activity changes were independent of changes in striatal dopamine levels.

Effects of polychlorinated biphenyls (PCBs) on brain tyrosine hydroxylase activity and dopamine synthesis in rats.

Literature reports suggest that polychlorinated biphenyls (PCBs) may alter dopaminergic neurotransmission in mammalian forebrain. In vitro, PCBs can decrease dopamine levels in PC 12 cells and studies of the structure-activity relationship (SAR) indicate that ortho-substituted (non-coplanar) PCB congeners are more active than para-substituted (coplanar) congeners. This report tested the hypothesis that ortho-substituted PCBs can selectively (vs para-substituted congeners) decrease dopamine synthesis in mammalian forebrain by inhibiting the activity of tyrosine hydroxylase, the rate-limiting enzyme in dopamine biosynthesis. In vitro effects of individual PCB congeners on activity of striatal tyrosine hydroxylase from two different rat strains were assessed. It was found that certain ortho-substituted PCB congeners (e.g., 2,2'-DCB) can inhibit tyrosine hydroxylase activity and dopamine synthesis by nearly 40% in minces of corpus striatum prepared from Sprague-Dawley and Long-Evans hooded rats. Comparatively, the ortho, meta-substituted PCB congener 2,2',5,5'-TeCB inhibited tyrosine hydroxylase activity only in striatal minces obtained from Sprague-Dawley rats, suggesting that genetic factors may influence the susceptibility of mammals to effects of PCBs that compromise brain dopamine synthesis. The PCB-induced inhibition of tyrosine hydroxylase activity in mammalian forebrain observed here appears to occur through indirect and as yet unknown mechanisms.

2-Phenylaminoadenosine stimulates dopamine synthesis in rat forebrain in vitro and in vivo via adenosine A2 receptors.

The adenosine agonist 2-phenylaminoadenosine (PAD) stimulated tyrosine hydroxylase activity in rat striatum in vitro. This effect was selectively blocked by the A2 antagonist 8-chlorostyrylcaffeine (CSC), suggesting an A2 receptor-mediated mechanism. PAD also produced a corresponding increase in striatal adenylyl cyclase activity. Using an in vivo model that measures presynaptic effects of drugs at dopamine nerve terminals, intracerebroventricular administration of PAD to rats stimulated tyrosine hydroxylase activity in striatum in a manner that was selectively blocked by CSC. These results suggest that PAD stimulates adenylyl cyclase and tyrosine hydroxylase activity, with a corresponding increase in dopamine synthesis, by activation of presynaptic A2-type receptors in mammalian forebrain.

A prospective, placebo-controlled, randomized trial of intravenous streptokinase and angioplasty versus lone angioplasty therapy of acute myocardial infarction.

BACKGROUND: The value of routine administration of intravenous thrombolytic agents during percutaneous transluminal coronary angioplasty (PTCA) therapy of acute myocardial infarction (MI) has not been determined. Therefore, we prospectively randomized 122 patients with evolving MI to PTCA therapy with or without adjunctive intravenous streptokinase therapy. METHODS AND RESULTS: Patients with ECG ST segment elevation who presented within 4 hours of symptom onset, had no contraindication to thrombolytic therapy, and were not in cardiogenic shock were enrolled. They were treated immediately with intravenous heparin (10,000 units) and oral aspirin (325 mg) and randomized to treatment with placebo or streptokinase (1.5 M units) administered intravenously over 30 minutes. Patients then were taken immediately to the catheterization laboratory, and those with suitable coronary anatomy underwent immediate PTCA. Subsequent clinical course, serial radionuclide ventriculography, and 6-month repeat angiography were analyzed. A total of 106 patients were treated with PTCA. Use of PTCA was similar for placebo (92%) and streptokinase (83%) groups. Angioplasty was successful in 95% of patients, with no difference in placebo (93%) and streptokinase (98%) groups. Serial radionuclide ventriculography demonstrated no difference in 24-hour (52 +/- 12% versus 50 +/- 12%) or 6-week (51 +/- 12% versus 51 +/- 13%) ejection fraction values for placebo and streptokinase groups, respectively. Contrast ventriculography demonstrated improvement in immediate (54 +/- 12%) versus 6-month (60 +/- 15%, p < 0.05) values for the overall group. No differences in 6-month values were present (58 +/- 15% versus 62 +/- 15%, p = NS) for placebo and streptokinase groups, respectively. Coronary angiography was performed in 75% of the 90 patients eligible for restudy. Arterial patency was 87% at 6 months, and coronary restenosis was present in 38% of patients. No differences in chronic patency or restenosis were detected for the two treatment groups. Although adjunctive intravenous streptokinase therapy did not improve outcome, it did complicate the hospital course. Hospitalization was longer (9.3 +/- 5.0 versus 7.7 +/- 4.4 days, p = 0.046) and more costly ($25,191 +/- 15,368 versus $19,643 +/- 7,250, p < 0.02). Transfusion rate was higher (39% versus 8%, p = 0.0001) and need for emergency coronary bypass surgery was greater (10.3% versus 1.6%, p = 0.03) for the streptokinase-treated patients. CONCLUSIONS: Adjunctive intravenous streptokinase therapy does not enhance early preservation of ventricular function, improve arterial patency rates, or lower restenosis rates after PTCA therapy of acute MI. Hospital course is longer, more expensive, and more complicated. For these reasons, PTCA therapy of acute MI should not be routinely performed with adjunctive intravenous streptokinase therapy.