Cough provocation tests: their clinical value.

Cough provocation testing was originally developed for testing new anti-tussive agents and still remains a mandatory tool in their development. The methodology has been developed into single breath and multiple breath techniques. Neither has been fully validated; however, single breath techniques have been used in a number of patient series of more than 100 patients. These studies show that the cough reflex is more sensitive in patients with cough and becomes normal on recovery. Studies in the common cold and in angiotensin converting enzyme inhibitor-associated cough suggest that this may be cause and effect. These results point to a utility for cough provocation testing in epidemiology but not in routine clinical practice. The one exception in clinical practice is in the assessment of patient post stroke where it is a better method, than voluntary cough testing, for assessing damage to the cough reflex. Other than use in epidemiology it remains an invaluable tool in hypothesis testing.

Cytotoxic and tubulin-interactive hemiasterlins from Auletta sp. and Siphonochalina spp. sponges.

Chemical and biological investigations of extracts from the sponge genus Auletta and two collections of Siphonochalina sp. have shown these organisms to be producers of the potent hemiasterlin class of antitumor agents. In addition to the previously known hemiasterlin (1) and hemiasterlin A (2), a new analogue, hemiasterlin C (3), was isolated and identified. The structures of 1 and 2 were assigned based on comparison to literature values, and 3 was identified on the basis of 1H NMR, 13C NMR, COSY, HSQC, and HMBC experiments. The cytotoxic and antitubulin activities of 1-3 were evaluated. In a comparative assay for inhibition of tubulin polymerization, the hemiasterlins were more potent than dolastatin 15 and equipotent with cryptophycin 1, but were somewhat less potent than dolastatin 10.

Vismiaphenones D-G, new prenylated benzophenones from Vismia cayennensis.

Following anti-HIV bioassay-guided fractionation, four new prenylated benzophenones, vismiaphenones D-G (7-10), were isolated from extracts of leaves of Vismia cayennensis. The structures were elucidated by spectral analyses. Only vismiaphenone D (7) exhibited HIV-inhibitory activity in the NCI primary screen.

Guttiferone F, the first prenylated benzophenone from Allanblackia stuhlmannii.

The HIV-inhibitory activity in extracts of Allanblackia stuhlmannii was tracked, via bioassay-guided fractionation, to a new member of the camboginol/guttiferone class of prenylated benzophenones, guttiferone F (1). The structure was solved by extensive NMR analyses and by acid-catalyzed conversion to 30-epi-cambogin (4). This is the first report of this compound type in the genus Allanblackia.

Pyranocoumarins from tropical species of the genus Calophyllum: a chemotaxonomic study of extracts in the National Cancer Institute collection.

(+)-Calanolide A, a novel dipyranocoumarin from the Malesian tree Calophyllum lanigerum var. austrocoriaceum, and a closely related compound, (-)-calanolide B, isolated from Calophyllum teysmannii var. inophylloide, are representatives of a distinct class of nonnucleoside HIV-1 specific reverse-transcriptase inhibitor under development as an AIDS chemotherapeutic. NCI repository specimens totalling 315 organic extracts from 31 taxa of Calophyllum were analyzed for related pyranocoumarins using a simple TLC system. A total of 127 extracts was initially classified as "positive"; eight out of the 31 taxa examined, representing perhaps 28 species already described (1/7-1/8 of all the species in this genus), contained prenylated coumarins, suggesting that these compounds, while sometimes abundantly present, are not widespread in the genus. Representative members of the TLC-positive extracts were partitioned between CH2C12 and 25% aqueous MeOH; the CH2C12-soluble materials were then analyzed by TLC and 1H NMR to confirm the presence of pyranocoumarins. The anti-HIV activity of the partitioned extracts are also presented. This study suggested that there are several distinctive coumarin chemotaxonomic markers distinguishing species of this genus.

The selective norepinephrine reuptake inhibitor, LY368975, reduces food consumption in animal models of feeding.

The compound, LY368975 ((R)-thionisoxetine) is a potent and selective inhibitor of the norepinephrine (NE) reuptake site. We evaluated the in vivo properties of LY368975 in various animal models. In mice, LY368975 prevented heart NE depletion by 6-hydroxydopamine with an ED50 of 1.22 mg/kg. In rats, orally administered LY368975 inhibited 3H-NE uptake into hypothalamic synaptosomes ex vivo with an ED50 of 2.5 mg/kg and 3H-tomoxetine binding to the NE transporter with an ED50 of 2.7 mg/kg. When rats were deprived of food for 18 hr, 10 mg/kg LY368975 was able to suppress food intake 1, 2 and 4 hr after reintroduction of the feed. In nonfasted rats trained to drink sweetened condensed milk, LY368975 produced a dose-dependent reduction in consumption with a 44% decrease at 3 mg/kg. At doses up to 10 mg/kg p.o., LY368975 produced no significant effects on locomotor activity suggesting the compound does not activate or sedate the animals at pharmacologically relevant doses. Therefore, LY368975 is an orally available and centrally active NE reuptake inhibitor that is capable of reducing food consumption in rodents. Compounds of this class may have use in the treatment of obesity and eating disorders.

Vasopressin/serotonin interactions in the anterior hypothalamus control aggressive behavior in golden hamsters.

Studies in several species of rodents show that arginine vasopressin (AVP) acting through a V1A receptor facilitates offensive aggression, i.e., the initiation of attacks and bites, whereas serotonin (5-HT) acting through a 5-HT1B receptor inhibits aggressive responding. One area of the CNS that seems critical for the organization of aggressive behavior is the basolateral hypothalamus, particularly the anterior hypothalamic region. The present studies examine the neuroanatomical and neurochemical interaction between AVP and 5-HT at the level of the anterior hypothalamus (AH) in the control of offensive aggression in Syrian golden hamsters. First, specific V1A and 5-HT1B binding sites in the AH are shown by in vitro receptor autoradiography. The binding for each neurotransmitter colocalizes with a dense field of immunoreactive AVP and 5-HT fibers and putative terminals. Putative 5-HT synapses on AVP neurons in the area of the AH are identified by double-staining immunocytochemistry and laser scanning confocal microscopy. These morphological data predispose a functional interaction between AVP and 5-HT at the level of the AH. When tested for offensive aggression in a resident/intruder paradigm, resident hamsters treated with fluoxetine, a selective 5-HT reuptake inhibitor, have significantly longer latencies to bite and bite fewer times than vehicle-treated controls. Conversely, AVP microinjections into the AH significantly shorten the latency to bite and increase biting attacks. The action of microinjected AVP to increase offensive aggression is blocked by the pretreatment of hamsters with fluoxetine. These data suggest that 5-HT inhibits fighting, in part, by antagonizing the aggression-promoting action of the AVP system.

Persistent spontaneous oral dyskinesias in haloperidol-withdrawn rats neonatally lesioned with 6-hydroxydopamine: absence of an association with the Bmax for [3H]raclopride binding to neostriatal homogenates.

To investigate the influence of dopamine (DA) nerves on haloperidol (HAL)-induced oral dyskinesias, rats were first injected at 3 days after birth with 6-hydroxydopamine HBr (200 micrograms i.c.v., salt form; 6-OHDA) or vehicle, after desipramine HCl (20 mg/kg i.p., 1 hr) pretreatment. Two months later HAL (1.5 mg/kg/day, 2 days a week for 4 weeks, then daily for 10 months) was added to the drinking water of half the rats. Numbers of vacuous chewing movements, recorded in 1-min increments every 10 min for 1 hr, increased from < 5 to about 17 oral movements per session in intact rats, 14 weeks after instituting HAL (P < .01 vs. intact rats drinking tap water). In HAL-treated 6-OHDA-lesioned rats, oral activity increased to > 30 oral movements per session (P < .01 vs. HAL-treated intact rats). These levels of oral activity persisted in intact and 6-OHDA-lesioned rats as long as HAL was administered. After 11 months of HAL treatment, but 8 or 9 days after HAL withdrawal, DA was found to be reduced 97%, whereas serotonin was increased 29% in the striatum of 6-OHDA-lesioned rats. In HAL-treated intact and lesioned rats the Bmax for DA D2 binding sites was elevated about 70%. With reverse transcription polymerase chain reaction, the mRNA level for DA D2L but not D2S receptors was also found to be elevated about 70%. In a fraction of 6-OHDA-lesioned rats that were observed for 8 months after HAL withdrawal, oral activity persisted without decrement and was not accompanied by a change in the Bmax or mRNA level for DA D2 receptors. These findings demonstrate that in rats largely DA-denervated as neonates, long-term HAL treatment produces an unusually high number of oral movements that persists for 8 months after HAL withdrawal and is not accompanied by an increase in DA D2 receptor expression.

Fluoxetine increases norepinephrine release in rat hypothalamus as measured by tissue levels of MHPG-SO4 and microdialysis in conscious rats.

The selective serotonin uptake inhibitor fluoxetine (10 mg/kg i.p.) increased tissue levels of the norepinephrine metabolite 3-methoxy-4-hydroxyphenylethylene glycol sulfate (MHPG-SO4) in rat hypothalamus, indicating an increased release of norepinephrine. Microdialysis studies in conscious rats showed that fluoxetine (10 mg/kg i.p.) increased extracellular concentrations of norepinephrine as well as serotonin in the hypothalamus. In contrast, desipramine (10 mg/kg i.p.) increased extracellular concentration of norepinephrine but not serotonin in the hypothalamus. Consistent with its mechanism of being a selective serotonin uptake inhibitor, local perfusion of fluoxetine (10 microM) caused a 7-fold increase in hypothalamic extracellular serotonin and a small non-significant increase in extracellular norepinephrine. The subsequent systemic injection of fluoxetine (10 mg/kg s.c.) after local perfusion caused a 3-fold increase in extracellular norepinephrine, indicating that fluoxetine's action leading to an increase in extracellular norepinephrine was not occurring in the terminal areas of the hypothalamus but elsewhere in the brain, possibly cell bodies in the locus coeruleus.

Structure-activity modifications of the HIV-1 inhibitors (+)-calanolide A and (-)-calanolide B.

The delta 7,8 olefinic linkages within (+)-calanolide A(1) and (-)-calanolide B(2) were catalytically reduced to determine impact on the anti-HIV activity of the parent compounds. In addition, a series of structure modifications of the C-12 hydroxyl group in (-)-calanolide B was made to investigate the importance of that substituent to the HIV-1 inhibitory activity of these coumarins. A total of 14 analogs were isolated or prepared and compared to (+)-calanolide A and (-)-calanolide B in the NCI primary anti-HIV assay. While none of the compounds showed activity superior to the two unmodified leads, some structure-activity requirements were apparent from the relative anti-HIV potencies of the various analogs.

Fluoxetine effects on serotonin function and aggressive behavior.

Fluoxetine inhibits serotonin uptake selectively and increases extracellular concentrations of serotonin in brain regions. The enhanced serotonergic neurotransmission resulting from increased action of that extracellular serotonin on postsynaptic receptors on target neurons results in various functional changes, reflecting the wide distribution of serotonin nerve terminals in brain regions that regulate numerous physiological functions. One consequence of fluoxetine administration in animals is a reduction of aggressive behavior, consistent with a larger body of data implicating serotonin as an important neurotransmitter modulator of aggression. In humans, preliminary data suggest that fluoxetine may also decrease aggressive behavior and feelings of anger or hostility. Further investigation of the potential usefulness of fluoxetine and other drugs that increase serotonergic function as a means of reducing anger, hostility, and aggressive behavior seems warranted.

Involvement of 5-HT2A receptors in the elevation of rat serum corticosterone concentrations by quipazine and MK-212.

The possible involvement of 5-HT2A or 5-HT2C receptors in the elevation of serum corticosterone in rats by quipazine (2-(1-piperazinyl)quinoline maleate) and MK-212 (6-chloro-(1-piperazinyl)pyrazine), direct-acting 5-HT receptor agonists, was investigated by the use of two newly available receptor antagonists, SB 200646A (N-(1-methyl-5-indolyl)-N'-(3-pyridyl)urea) and MDL 100,907 (R-(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]- 4-piperidinemethanol). MDL 100,907 blocked the increase in serum corticosterone elicited by quipazine and MK-212 with ED50 values of 0.0028 and 0.0027 mg/kg, s.c., respectively. In contrast, SB 200646A only partially antagonized the serum corticosterone concentration increases by quipazine and MK-212 even at the highest dose tested, 40 mg/kg, i.p. Because published data show the affinities of MDL 100,907 and SB 200646A for 5-HT2C receptors to be nearly identical, whereas the affinity of MDL 100,907 for 5-HT2A receptors is 17500-fold higher than that of SB 200646A, our findings suggest that 5-HT2A receptors rather than 5-HT2C receptors mediate the serum corticosterone increases by both quipazine and MK-212.

New pyranocoumarins isolated from Calophyllum lanigerum and Calophyllum teysmannii.

During a chemotaxonomic survey of Calophyllum extracts present in the National Cancer Institute's natural product repository, four new pyranocoumarins were isolated from extracts of C. lanigerum var. austrocoriaceum and C. teysmannii var. inophylloide (King.) P. F. Stevens (Clusiaceae). The structure elucidation and anti-HIV activity of calanolide E2 (4), cordatolide E (5), pseudocordatolide C (6), and calanolide F (9), along with a simple prenylated coumarin precursor (11), are described here.

On the in-vivo modulation of neostriatal dopamine release by fluoxetine and 5-hydroxy-L-tryptophan in conscious rats.

To help determine the nature of serotonergic regulation of dopamine activity in the brain an in-vivo microdialysis study has been performed in conscious rats to investigate the modulation of dopamine release in the neostriatum by 5-hydroxytryptamine (5-HT). The 5-HT uptake inhibitor, fluoxetine, and the 5-HT precursor, 5-hydroxy-L-tryptophan (5-HTP), were used to produce an increase in extracellular 5-HT concentration Systemic administration of fluoxetine (10 mg kg-1, s.c.) produced a 2- to 3-fold increase in extracellular 5-HT concentration but did not change extracellular dopamine concentration in the neostriatum. Co-administration of fluoxetine and 5-HTP (40 mg kg-1, s.c.; 60-90 min after fluoxetine) caused a highly significant tenfold increase in extracellular 5-HT concentration in the neostriatum with a slight but non-significant decrease in extracellular dopamine concentration. Pergolide, a dopamine D2 agonist, given systemically caused a dramatic decrease in extracellular dopamine concentration demonstrating the responsiveness of the neurons. These results demonstrate that high concentrations of extracellular 5-HT do not modulate dopamine release in the neostriatum. The possibility that different 5-HT receptor subtypes may mediate different regulation of dopamine release remains to be explored.

The asthma death problem revisited.

1. Asthma is a potentially dangerous disease. Asthma deaths occur at varying degrees throughout the world and there is evidence for epidemics occurring in different populations at different times. Such epidemics have been noted as early as last century. Much investigation has been made into the role of inhaled beta-adrenoceptor agonists in these epidemics and indeed there is some evidence fenoterol is implicated. However, evidence for other beta-adrenoceptor agonists is not substantiated. 2. It is also noted that asthma deaths are not all the same, there being at least two types: deaths occurring in chronic severe asthmatics related to disease severity and sudden deaths of an anaphylactic nature that can occur in asthmatics of any disease severity. 3. The important next step in the study of asthma deaths is to concentrate on other important factors rather than the role of therapy which cannot explain all the epidemics and the background instance of asthma death.

A comparison of the effects of an alpha-agonist, an anti-muscarinic agent and placebo on intranasal histamine challenge in allergic rhinitis.

Autonomic receptors play a part in the physiology and pathology of the nasal mucosa. The effect of an alpha-agonist and an anti-muscarinic agent on histamine-challenge was examined on patients with perennial allergic rhinitis. Nine patients received saline, oxitropium bromide 0.075%, or xylometazoline hydrochloride 0.1% in a double-blind fashion. Sequential challenge with increasing doses of histamine were given and resistance changes, sneezes and volume and content of secretion measured. Histamine challenge produced dose-related increases in nasal resistance (P < 0.0001), lavage fluid volume (P < 0.01) and total protein (P < 0.01). Following xylometazoline, histamine produced little increase in resistance compared with saline and oxitropium bromide (P < 0.0001). The latter reduced the dose-related increase in resistance (P < 0.01) and nasal lavage fluid volume (P = 0.0007) and total protein (P = 0.023) seen with saline. These results confirm the importance of alpha-adrenergic and muscarinic receptors in the human nasal mucosa and suggest mechanisms of action for these drugs in perennial allergic rhinitis.

Pharmacokinetics of intravenous fluticasone propionate in healthy subjects.

1. Fluticasone propionate (FP) is a potent glucocorticoid used in the treatment of asthma. Prior to reporting the pharmacokinetics following the inhaled and oral routes, the pharmacokinetics need to be established following intravenous dosing. The present study determines the intravenous pharmacokinetics of FP, using non-compartmental analysis, in healthy male subjects over the 250 to 1000 micrograms dose range. 2. The pharmacokinetics of FP can be regarded as being linear over this dosing range. FP was extensively distributed within the body (Vss 3181), rapidly cleared (CL 1.1 l min-1) with a terminal elimination half-life of 7.8 h and a mean residence time of 4.9 h. 3. In order that future pharmacokinetic/pharmacodynamic and other modelling can be carried out, the plasma concentration-time profiles were parameterized using a model based on sums of exponentials, the appropriateness of this model was justified as the secondary kinetic parameters from the model were similar to those obtained using non-compartmental analysis.

Capsaicin cough sensitivity increases during upper respiratory infection.

The mechanism of cough associated with upper respiratory infection (URI) is poorly understood. This paper reports a study of the role of altered sensitivity of capsaicin-sensitive airway nerves. In a prospective study, baseline (B) capsaicin-induced cough and methacholine-induced airway responsiveness were measured in 103 healthy volunteers. During the following year, 31 subjects reattended for challenge testing during URI (I) and after recovery (R). The log concentration of capsaicin required to elicit two coughs (C2) was significantly lower during infection than recovery but not baseline [median (interquartile range) B = 0.59 (0.28-1.20), I = 0.27 (0-0.89), R = 0.89 (0.28-1.49)]. Log C5 (concentration causing five coughs) was lower during infection than baseline and recovery [B = 1.79 (1.20-2.70), I = 1.49 (0.89-2.08), R = 1.79 (1.20-2.40)]. FEV1 and PC15 methacholine values were unchanged during infection compared to baseline. Subjects with dry cough (n = 14) had lower C5 values during infection than both baseline and recovery, and lower C2 values during infection than recovery; in these subjects, increase in capsaicin sensitivity correlated with cough severity score. Subjects with productive cough or no cough showed no consistent changes during infection. Twenty-six control subjects who reattended without URI showed no change in capsaicin sensitivity. Upper respiratory infection may cause cough as a result of increased sensitivity of capsaicin-sensitive afferent airway nerves without affecting airway calibre or responsiveness.