Suitability of botanical extracts as components of complex mixtures used in herbal tea infusions-challenges and opportunities.

Herbal tea is a mainstay dosage form in practically all systems of traditional medicine and widely used in modern alternative and complementary medicine. Incorporating botanical extracts into herbal tea formulations is of vital interest to manufacturers as it allows for the use of herbal ingredients that would otherwise not be suitable for the dosage form, for instance, dosing requirements, solubility in water, sensory constraints etc. Furthermore, reducing the amount of ingredients in a formula increases compliance with dosing recommendations and thus therapeutic benefit. However, formulating with botanical extracts comes with challenges, ranging from sourcing ingredients of appropriate quality, developing suitable methods for quality control with combinations of (herbal) ingredients, processing constraints such as hygroscopicity, solubility, dispersibility, homogeneity of distribution, and packaging machinability, all the way to stability required for hot-water infusion. We report on experiences with overcoming such challenges in a set of examples and provide guidance to the extract industry on how to tap into the bagged tea sector with better suited or tailor-made solutions for the formulator.

Blunted response to low oxygen of rat respiratory network after perinatal ethanol exposure: involvement of inhibitory control.

Acute ethanol depresses respiration, but little is known about chronic ethanol exposure during gestation and breathing, while the deleterious effects of ethanol on CNS development have been clearly described. In a recent study we demonstrated that pre- and postnatal ethanol exposure induced low minute ventilation in juvenile rats. The present study analysed in juvenile rats the respiratory response to hypoxia in vivo by plethysmography and the phrenic (Phr) nerve response to ischaemia in situ. Glycinergic neurotransmission was assessed in situ with strychnine application and [(3)H]strychnine binding experiments performed in the medulla. After chronic ethanol exposure, hyperventilation during hypoxia was blunted in vivo. In situ Phr nerve response to ischaemia was also impaired, while gasping activity occurred earlier and recovery was delayed. Strychnine applications in situ (0.05-0.5 microM) demonstrated a higher sensitivity of expiratory duration in ethanol-exposed animals compared to control animals. Moreover, [(3)H]strychnine binding density was increased after ethanol and was associated with higher affinity. Furthermore, 0.2 microM strychnine in ethanol-exposed animals restored the low basal Phr nerve frequency, but also the Phr nerve response to ischaemia and the time to recovery, while gasping activity appeared even earlier with a higher frequency. Polycythaemia was present after ethanol exposure whereas lung and heart weights were not altered. We conclude that chronic ethanol exposure during rat brain development (i) induced polycythaemia to compensate for low minute ventilation at rest; (ii) impaired the respiratory network adaptive response to low oxygen because of an increase in central glycinergic tonic inhibitions, and (iii) did not affect gasping mechanisms. We suggest that ethanol exposure during early life can be a risk factor for the newborn respiratory adaptive mechanisms to a low oxygen environment.

A haplotype of the DRD1 gene is associated with alcohol dependence.

BACKGROUND: The D1 dopamine receptor has been involved in a number of brain functions, including motor control, inattentive symptoms and reward and reinforcement mechanisms. Indeed, DRD1 antagonists may reduce cocaine-seeking behavior and the acquisition of cocaine-cue associations. The D1.1/r4532 marker of the DRD1 gene has been associated with a large set of phenotypes including addictive behaviors, but none with alcohol dependence per se. METHODS: We analyzed a population of 134 patients with alcohol dependence, also assessing more homogeneous (severe) phenotypes, comparing this sample with a healthy control population, assessing two SNPs within the DRD1 gene in order to depict the role of DRD1 polymorphisms and haplotypes. RESULTS: The T allele of the rs686 polymorphism within DRD1 gene was significantly more frequent in patients with alcohol dependence (p = 0.0008), with a larger excess for patients with severe dependence (p = 6 x 10(-6)), and even more for patients with severe complications such as withdrawal seizures (p = 7 x 10(-7)). A specific haplotype rs686*T-rs4532*G within the DRD1 gene was significantly more precisely associated with alcohol dependence in our sample (p = 5 x 10(-6)). CONCLUSIONS: Even though chance finding cannot be ruled out, convergent evidence is given that the DRD1 gene is a susceptibility gene in alcohol dependence, regarding the fact that relying on more homogeneous phenotypes (i.e., more severe patients) and more informative genetic markers (i.e., haplotypes) reinforce the initial association.

Circling behaviour in rats with unilateral lesions of the nigrostriatum induced by 6-hydroxydopamine: changes induced by oral administration of cytidine-5'-diphosphocholine.

A unilateral administration of 6-hydroxydopamine into the nigrostriatal system of the rat was responsible for ipsiversive circling behaviour in response to administration and, in time, of contraversive circling behaviour in response to L-DOPA and apomorphine. This contraversive circling behaviour appears to be mediated by the development, on the lesioned side, of supersensitivity of postsynaptic dopamine receptors. Subchronic treatment with cytidine-5'-diphosphocholine (p.o.) by itself was devoid of behavioural effects. The CDP-choline did not modify the apomorphine-induced stimulant effect but potentiated the circling behaviour produced by L-DOPA and amphetamine. The data show that the effects of CDP-choline were mediated by a presynaptic mechanism: the potentiation of the effects of L-DOPA cannot be explained by an activation of tyrosine hydroxylase, but seems to be related to an improvement of release of newly synthesized dopamine from exogenous L-DOPA.

Absence of the adenosine A(2A) receptor or its chronic blockade decrease ethanol withdrawal-induced seizures in mice.

Considering the existing interactions between ethanol and adenosine, the influence of the genetic impairment of the adenosine A(2A) receptor has been examined upon the seizures occurring at the cessation of chronic ethanol intake or 'ethanol withdrawal' in male mice. Acute clearance of ethanol did not differ between adenosine A(2A) receptor knockout and wild-type mice. Mice were exposed for 10 days to a diet consisting of a milky chocolate drink that contained increasing concentrations (1.8, 3.6 and 6.3% v/v) of ethanol. Adenosine A(2A) receptor knockout mice ingested similar amounts of the fluid, either containing alcohol or not, as did the controls. The severity of handling-induced convulsions during withdrawal was significantly reduced in the adenosine A(2A) receptor knockout mice as compared with their wild-type controls. The selective adenosine A(2A) receptor antagonist ZM 241385 (20 mg/kg) also significantly attenuated the intensity of withdrawal-induced seizures occurring in wild-type male mice when intraperitoneally administered twice daily during the last 5 days of the forced alcohol intake. These results suggest that selective adenosine A(2A) receptor antagonists may be useful in the treatment of alcohol withdrawal.

Cyamemazine decreases ethanol intake in rats and convulsions during ethanol withdrawal syndrome in mice.

The effect of cyamemazine a dopamine D2 receptor antagonist on voluntary ethanol consumption in rats and on ethanol withdrawal in mice was examined. Male Sprague-Dawley rats were tested in a free choice (water and 10% ethanol) experiment and consumed 5 g/kg ethanol daily. Rats were treated daily IP with cyamemazine (0.5, 1, or 2 mg/kg) or acamprosate (100 mg/kg) during 2 weeks. Both acamprosate and 1 mg/kg cyamemazine significantly decreased ethanol intake by 45% without affecting either fluid or food intake. The lowest dose of cyamemazine had no effect on alcohol intake but increased food intake. The highest dose had no effect on any variables. During the post-treatment period, only 1 mg/kg cyamemazine decreased both ethanol and fluid intakes. Mice were made dependent on alcohol using a chocolate fluid diet containing increasing concentrations of alcohol and withdrawn after 9 days. Mice were treated with cyamemazine (1 or 0.5 mg/kg, respectively) or with the same doses of lorazepam acutely on the day of withdrawal or chronically (during alcohol treatment). Both chronic and acute cyamemazine and lorazepam treatments decreased convulsions during ethanol withdrawal. Both acute treatments decreased locomotor activity in control and alcohol dependent mice. Chronic treatment had no effect on locomotor activity. We suggest that cyamemazine could reduce alcohol consumption by antagonizing the activation of the dopaminergic pathways during the induction of alcohol dependence. The action of cyamemazine on 5-HT3 receptors could also explain its effect on alcohol convulsions during withdrawal convulsions.

Pre-exposure to alcohol does not sensitize to the rewarding effects of cocaine.

The conditioned place preference (CPP) induced by cocaine 2.5 mg/kg was measured in rats pre-exposed to ethanol (14 days with only 10% v/v ethanol followed by a free choice between ethanol solution and water for 14 days). Rats were divided according to their alcohol intake during the free choice period into low-drinking (<3 g/kg per day), intermediate-drinking and high-drinking (> 4 g/kg per day) rats. Cocaine-induced CPP was not modified in high-drinking rats relative to controls. Low-drinking rats had a lower CPP than high-drinking rats and controls. We conclude that pre-exposure to alcohol did not sensitize to the cocaine rewarding effects, and that alcohol low-drinking rats showed the lowest preference for cocaine.

Ethanol potentiates lipopolysaccharide- or interleukin-1 beta-induced nitric oxide generation in RBE4 cells.

Our present study investigated the effects of ethanol treatment on inducible nitric oxide (NO) synthase pathway from lipopolysaccharide- or interleukin-1 beta-treated cultured rat blood-brain barrier cell line (rat brain endothelial 4 cells: RBE4 cells). Cells were lipopolysaccharide- or interleukin-1 beta-treated with or without ethanol (50, 100 or 200 mM) for 16 or 24 h. Inducible NO synthase activity and mRNA expression were measured using Griess reaction and reverse transcription-polymerase chain reaction (RT-PCR) respectively. In the absence of lipopolysaccharide or interleukin-1 beta, ethanol treatments failed to stimulate inducible NO synthase gene expression. Lipopolysaccharide or interleukin-1 beta increased nitrite production and inducible NO synthase mRNA levels, and ethanol potentiated this effect. We concluded that ethanol could aggravate the consequences of NO generation by RBE4 cells after inducible NO synthase induction following inflammation or sepsis. This ethanol action on NO generation could contribute to circulatory failure associated with shock due to sepsis or hemorrhage, and alter blood-brain barrier permeability.

Experimental findings in the study of the reduction of alcohol intake.

Alcohol dependence represents a major problem in public health and different animal models of dependence have been developed in rodents with the aim of studying the mechanisms of alcohol abuse. Different ways of animal alcoholisation have been established. They permit a better understanding of which neurotransmitter system is involved in the regulation of alcohol dependence. Considerable attention has been given to the role of serotonin in the control of both alcohol craving and alcohol related pathologies, i.e. anxiety, aggression or memory loss. In conclusion, the use of animal models of alcohol abuse facilitates the understanding of alcohol behavior and permits the development of new therapeutic agents.

Ethanol intake and 3H-serotonin uptake I: A study in Fawn-Hooded rats.

Ethanol intake and synaptosomal 3H-serotonin uptake were studied in male Fawn-Hooded and Sprague-Dawley rats. Fawn-Hooded rats consumed more alcohol and more water than Sprague-Dawley rats. Plasma alcohol levels of Sprague-Dawley rats were not detectable but were about 5 mg/dl in Fawn-Hooded rats. Ethanol intake increased the Vmax of serotonin uptake in Fawn-Hooded rats in hippocampus and cortex, but not in thalamus. In Fawn-Hooded rats, serotonin uptake (Vmax) was higher than in Sprague-Dawley rats cortex. Ethanol intake reduced the Vmax of serotonin uptake in Fawn-Hooded rats in hippocampus and cortex. In cortex, the carrier affinity for serotonin was increased in alcoholized Fawn-Hooded rats. These results indicate that synaptosomal 3H-serotonin uptake is affected by ethanol intake. In Fawn-Hooded rats, high ethanol consumption is associated with high serotonin uptake. In rats presenting high serotonin uptake, alcoholisation reduces 3H-serotonin internalisation in synaptosomes, indicating a specific sensitivity to alcohol intake of serotonin uptake system.

Ethanol intake and 3H-serotonin uptake. II: A study in alcoholic patients using platelets 3H-paroxetine binding.

The kinetic parameters of 3H-paroxetine binding and 3H-serotonin uptake were studied in platelets of alcoholic patients. There was no difference between alcoholic and non alcoholic subjects in 3H-paroxetine binding. When binding and 3H-serotonin uptake were studied, in the same plasma of the same subjects, the Vmax of serotonin uptake was increased in alcoholics. The data confirm the involvement of serotonin uptake system in alcohol dependence and suggest that serotonin uptake and paroxetine binding sites may be regulated independently in this pathology.

Platelets 3H 5-HT uptake in descendants from alcoholic patients: a potential risk factor for alcohol dependence?

We studied platelet 3H-serotonin uptake in 32 former alcoholics, withdrawn for from 1 month to 22 years, in their descendants (21.7 +/- 1.6 years old, n = 17; 10.9 +/- 0.7 years old; n = 19), and in respective control groups, paired in age and sex. All of the alcoholics presented high 3H-serotonin uptake (Vmax = 10.88 +/- 4.23 pmoles/10(8)pl/30 sec., vs. 0.93 +/- 0.15 pmole/10(8)pl/30 sec. Their descendants also showed high platelet serotonin uptake: 3.94 +/- 1.44 pmoles/10(8)pl/30 sec., vs. 0.93 +/- 0.15 pmoles/10(8)pl/30 sec. for adult descendants, and 5.99 +/- 2.23 vs. 0.84 +/- 0.15 pmole/10(8)pl/30 sec. for young descendants. All subjects were free of alcoholisation (biological parameters studied were blood ethanol concentration, gamma glutamyl transferase and mean corpuscular volume), and dependence of former alcoholics was evaluate by using, a posteriori, the CAGE test. In descendants, 28% of the subjects have Vmax values higher than the highest of the control group. Alcohol, in vitro, (54 mM) did not affect serotonin uptake in any group. These results indicate that in descendants of alcoholics, platelet serotonin uptake is altered, without modification of sensitivity to ethanol. The genetic basis of alcohol dependence could be linked with the platelet serotonin transport.

Alcohol and GABA: ethanol intake modifies hippocampal nipecotic acid binding in ethanol-preferring and non-preferring rats.

3H-nipecotic acid (3H-NIP) binding to GABA uptake recognition sites was studied in the hippocampus of 3 groups of male, Long Evans rats: Group 1: ethanol-naive rats (ENR); Group II: ethanol-preferring rats (DR) and non-preferring rats (NDR), which had consumed about 5 g.kg-1.d-1 and 1 g-1.d-1 of alcohol respectively in the form of a 12% ethanol solution prior to 3H-NIP binding analysis; Group III: DR and NDR who had had no access to ethanol for 21 d after the initial exposure of ethanol solution (28 d). Binding studies showed that ethanol drunk by both DR and NDR in Group II decreased 3H-NIP binding (Bmax decreased) with an enhancement of affinity (KD decreased). In rats subjected to withdrawal of ethanol (Group III), affinity of 3H-NIP for GABA uptake sites was higher than in controls (Group I), but lower than in Group II, Bmax in this group being higher than in the 2 other groups. In Group III, KD was higher in DR than in NDR. These results showed that ethanol intake, in a free-choice paradigm, altered 3H-NIP binding, and that differences in ethanol intake between DR and NDR were associated with differences in sensitivity of hippocampal GABA uptake sites. These differences in 3H-NIP binding could either precede ethanol intake, or be a direct result from it. The results, together with data from other laboratories suggest that: 1), 3H-NIP binding sites are involved in the regulation of ethanol intake; 2), 1 factor responsible for individual differences in ethanol response is reflected by the GABA uptake system.

Noradrenergic system: effect of DSP4 and FLA-57 on ethanol intake in ethanol preferring rats.

Ethanol preferring rats (male Long-Evans; n = 6) were selected as drinking rats (DR) and treated with DSP4 (50 mg.kg-1 IP) at the end of the preference selection. Two more groups received DSP4 (50 mg.kg-1 IP) + the inhibitor of dopamine beta-hydroxylase FLA-57 (1 mg.kg-1.d-1 during two weeks), IP (n = 5) or FLA-57 alone (1 mg.kg-1.d-1 during two weeks IP) (n = 5). The control DR group (n = 6) received NaCl 0.9%. 3H-Noradrenaline uptake was studied at the 17th day of treatment in DR, treated or not with DSP4, and in ethanol naive rats treated (n = 6) or not (n = 6) with DSP4 (50 mg.kg-1, IP) DSP4 does not modify ethanol intake in DR, and both treated groups (DR or ethanol naive rats). 3H-Noradrenaline uptake was decreased (about 60%), both in cortex and hippocampus. But the association of FLA-57 and DSP4 decreases both ethanol and fluid intakes. It was suggested 1) that the 40% of intact neurons was able to compensate the DSP4-induced noradrenergic neurons destruction, 2) that the destruction of noradrenergic pathways (FLA-57 + DSP4) is associated with a decrease in ethanol intake but also in fluid intakes, suggesting finally 3) that the modulation of ethanol intake by the noradrenergic system was partial or indirect.

Intracerebroventricular injection of antisense oligos to nNOS decreases rat ethanol intake.

Nitric oxide (NO) has been implicated in alcohol drinking behavior using NO synthase (NOS) inhibitors that are nonselective of the different isoforms of NOS. In the brain, there are two constitutive isoforms of NOS, neuronal NOS (nNOS) and endothelial NOS (eNOS). We used an antisense oligodeoxynucleotide directed against nNOS in ethanol dependent male Wistar rats to examine the specific contribution of nNOS in the control of ethanol intake. Rats were subjected to a free-choice situation water/ethanol (10% v/v) after chronic ethanol intoxication by inhalation of ethanol vapor. During the free-choice situation, rats were twice daily for 4 days intracerebroventricularly injected with either saline, or end-capped phosphorothioate-protected antisense or mismatch oligodeoxynucleotide (25 microg/4 microl per injection), or acamprosate (1 mg/kg body weight) as reference product for its anticraving properties. Our results showed that the antisense treatment, but not the mismatch treatment, reduced both ethanol intake and ethanol preference during treatment and posttreatment periods (by 25-30%) without alteration of the body weight gain. The antisense treatment, but not the mismatch treatment, also down-regulated nNOS mRNA levels (by 30%) and NOS activity in the hippocampus. The anticraving drug, acamprosate reduced both ethanol intake (by 58%) and ethanol preference. All these results suggest that nNOS is involved in the regulation of alcohol dependence.

Isolation and striatal (3H) serotonin uptake: role in the voluntary intake of ethanol by rats.

Ethanol preferring rats were selected and showed a constant voluntary intake of a 12 percent ethanol solution during 14 days (about 5 g/kg body weight daily). Analysis of 3H serotonin uptake by striatal synaptosomes showed that steady state 3H serotonin synaptosomal levels were lower in alcohol preferring rats. Grouping these rats (5 per cage) reduced both voluntary intake of ethanol and synaptosomal 3H serotonin uptake. Furthermore, blocking the serotonin uptake by clomipramine 5 mg X kg-1 or 10 mg X kg-1 also reduces voluntary intake of ethanol. These data are in agreement with the hypothesis of a modulation of the voluntary intake of ethanol both by chemical and housing stimulation of striatal receptors for serotonin.

Acamprosate modulates synaptosomal GABA transmission in chronically alcoholised rats.

Male Sprague-Dawley rats were pulmonary alcoholised for 30 days. Six were treated with acamprosate (400 mg/kg/day, PO) during alcoholisation. The control nonalcoholised group also received acamprosate (400 mg/kg/day, PO) during the 30 days. At the end of the experiment, brains areas (cortex, hippocampus, thalamus, striatum, and olfactory bulbs) were dissected for the study of synaptosomal 3H-GABA uptake. In another experiment, GABA levels were determined in the same areas using HPLC with electrochemical detection. Blood ethanol levels were also measured during alcoholisation. Acamprosate treatment did not modify blood ethanol levels. In cortex and olfactory bulbs, alcoholisation increased 3H-GABA uptake (Vmax) with an increase in the affinity (Km). 3H-GABA uptake was not affected by alcoholisation in other brain areas. In hippocampus and thalamus, acamprosate treatment enhanced 3H-GABA uptake (Vmax) only in alcoholised rats. Moreover, in thalamus, alcoholisation enhanced GABA levels. The effect of alcohol and acamprosate on GABA presynaptic events is discussed and it is concluded that the action of ethanol and acamprosate on GABA transport could be, in part, responsible for the modulation by acamprosate treatment of ethanol behaviour.

A homotaurine derivative reduces the voluntary intake of ethanol by rats: are cerebral GABA receptors involved?

The effects of some derivatives of homotaurine (3 APS), the well known GABA agonist, were tested on the voluntary intake of ethanol by rats. Spontaneously ethanol drinking rats (DR) were selected and had a constant voluntary intake of ethanol by rats. Spontaneously ethanol drinking rats (DR) were selected and had a constant voluntary intake of a 12% ethanol solution (VIE) during 14 days (about 5 g/kg body weight daily). Calcium acetylhomotaurine (0.26 and 0.52 mmol/kg daily IP) significantly reduced VIE and this was inhibited by the GABA antagonist bicuculline (2 mg/kg IP). The conditioned aversion test to saccharin was negative. Bicuculline alone did not affect VIE. Other homotaurine (3-APS) derivatives: sodium acetyl homotaurine (Na AOTA), homotaurine (OTA), sodium acetyltaurine (Na A TA) and calcium chloride (CaCl2) did not affect VIE. These data suggest that the gabaergic system could be implicated in VIE. MERAM Lab. patent.

A novel indication for transvenous lead extraction: upgrading implantable cardioverter defibrillator systems.

UNLABELLED: Technological advances have resulted in the development of dual chamber pacemaker/defibrillator systems with smaller pectoral 'active cans'. Patients now have the option of upgrading from abdominal to pectoral or from single to dual chamber devices. In addition, due to the potential complications which may arise with abandoned ICD leads, extraction of preexisting leads may be preferable. METHODS AND RESULTS: Twenty consecutive patients (11 males), underwent lead extraction and upgrade, either from an abdominal to a pectoral, or from a single to a dual chamber device. The mean age was 62+/-18 years and mean implant duration was 50+/-14 months. Indications for extraction included lead fracture/malfunction (13), ERI/EOL (2), new SVT/VT (2), long charge times (2), and impending erosion (1). An initial attempt was made to remove the lead with gentle traction. If excessive scar tissue prohibited extraction, then a laser sheath was employed. Reimplantation proceeded following standard protocol. Clinical success was achieved in all patients. Eleven of thirty leads were removed with traction. The remaining 19 leads required removal with the laser sheath. All ICD reimplants were placed in the left pectoral position, of which 10 were dual chamber. The mean defibrillation threshold was 9.5+/-5.8 Joules. There were no procedure related perforations or deaths. At follow up (13+/-10 mos. ) there were no infections, lead malfunctions or venous thromboses. There were two deaths, both from intractable heart failure. CONCLUSIONS: This study demonstrates that, when indicated, ICD leads can be safely extracted and systems successfully upgraded to take advantage of new technology.

Attenuation by antidepressant drugs of alcohol intake in rats.

Ethanol preferring rats were selected and showed a constant voluntary intake of a 12% ethanol solution during 14 days (about 5 g/kg body weight daily). These alcohol preferring rats were daily IP injected during two weeks with different antidepressant drugs, according to their specificity of action: nomifensine (5 and 10 mg/kg) and maprotiline (2.5 and 5 mg/kg) (dopamine uptake inhibitors), desipramine and metapramine (5 and 10 mg/kg) (noradrenaline uptake inhibitors) clomipramine and doxepin (5 and 10 mg/kg) (serotonin uptake inhibitors). Only desipramine, 5 and 10 mg/kg, metapramine, 10 mg/kg, clomipramine, 5 and 10 mg/kg and doxepin, 10 mg/kg, were able to reduce significantly the ethanol intake. These drugs specifically inhibit noradrenaline or serotonin uptake. These data lead us to think that norepinephrine and/or serotonin, but not dopamine, are involved in the voluntary intake of alcohol.