Celine H.K. Cheng

Angiofensin converting enzyme density is increased in temporal cortex from patients with Alzheimer's disease

The present study assesses the binding density of the selective angiotensin converting enzyme (ACE) radioligand [3H]ceranapril in brain tissue homogenates derived from patients with Alzheimers disease and those from age-, sex- and post-mortem delay-matched neurologically normal patients. Saturation studies with [3H]ceranapril identified that the specific binding (defined by captopril, 10 μM) was homogenous and of high affinity. ACE inhibitor recognition site density was higher by some 70% in the temporal cortex (Brodmann area 22) from Alzheimers patients whereas densities were similar in frontal cortex and cerebellum when compared to control tissue. It is unknown whether this apparently selective alteration in ACE density is directly related to. or a compensatory effect of the disease, but it provides additional support for the development of compounds which interact with the central angiotensin system as novel therapies for cognitive dysfunction.

Differential modulation of extracellular levels of 5-hydroxytryptamine in the rat frontal cortex by (R)- and (S)-zacopride.

1 The ability of various anxiolytic and potential anxiolytic agents to modify 5‐hydroxytryptamine (5‐HT) release in the frontal cortex of the rat was assessed by the microdialysis technique. 2 The benzodiazepine receptor agonist, diazepam (2.5 mg kg−1, i.p.), the 5‐HT1A receptor agonist 8‐hydroxy‐2‐(di‐n‐propylamino) tetralin (8‐OH‐DPAT, 0.32 mg kg−1, s.c.) and the 5‐HT1A receptor partial agonist buspirone (4.0 mg kg−1, i.p.) maximally reduced extracellular levels of 5‐HT in the rat frontal cortex by approximately 50–60%, 70–80% and 30–40%, respectively. 3 (R)‐zacopride (1.0–100 μg kg−1, i.p.) dose‐dependently reduced extracellular levels of 5‐HT in the rat frontal cortex (approximately 80% maximal reduction) whereas the other 5‐HT3 receptor antagonists ondansetron (10 μg kg−1, i.p.) and (S)‐zacopride (10–100 μg kg−1, i.p.) were ineffective. 4 In contrast to (S)‐zacopride (100 nm; administered via the microdialysis probe), (R)‐zacopride (1.0–100 nm; administered via the microdialysis probe) induced a concentration‐dependent reduction in extracellular levels of 5‐HT in the rat frontal cortex (approximately 70% maximal reduction). 5 In contrast to ondansetron (100 μg kg−1, i.p.), (S)‐zacopride (10–100 μg kg−1, i.p.) dose‐dependently reversed the (R)‐zacopride (10 μg kg−1, i.p.) induced reduction in extracellular levels of 5‐HT in the rat frontal cortex. The highest dose of (S)‐zacopride (100 μg kg−1, i.p.) completely prevented the (R)‐zacopride response. In addition, (S)‐zacopride (100 nm; administered via the microdialysis probe) attenuated the inhibitory action of (R)‐zacopride (10 nm; administered via the microdialysis probe) on extracellular levels of 5‐HT in the rat frontal cortex. 6 In conclusion, the present study provides further evidence of the ability of diazepam, 8‐OH‐DPAT and buspirone to reduce the activity of the central 5‐hydroxytryptaminergic system in vivo. Furthermore, the results indicate that the ability of (R)‐zacopride to reduce the in vivo release of 5‐HT in the rat frontal cortex does not correlate with its 5‐HT3 receptor antagonism. However, the differential affinity of (R)‐ and (S)‐zacopride for a (S)‐zacopride‐insensitive (R)‐zacopride site in rat cerebral cortex mirrors the relative activity of the two zacopride stereoisomers to modify the in vivo release of 5‐HT in the frontal cortex of the rat and their ability to release suppressed behaviour in animal models of anxiety.

The pharmacological characterization of 5-HT3 receptor binding sites in rabbit ileum: Comparison with those in rat ileum and rat brain ☆

Abstract We have further characterized the 5-HT 3 receptors in rat and rabbit tissues by evaluating the binding of the 5-HT 3 receptor ligand, [ 3 H]GR67330 to homogenates of rabbit ileum, rat ileum and rat brain (entorhinal cortex). In each tissue specific [ 3 H]GR67330 binding represented a single saturable, high affinity site ( K d = 0.14, 0.18, 0.076 nM in rabbit ileum, rat ileum and rat brain respectively). The densities of sites present in rabbit and rat ileum were similar to that present in rat brain ( B max = 63, 47, 72 fmol/mg protein in rabbit ileum, rat ileum and rat brain respectively). In each tissue, 5-HT 3 receptor agonists and antagonists potently competed for [ 3 H]GR67330 binding. Derived inhibition constants were similar in rat ileum and brain. However marked differences in IC 50 s were apparent for rabbit ileum compared with rat brain or ileum. These were most apparent with agonists. Thus, m CPBG [1-( meta -chlorophenylbiguanide)], phenylbiguanide, 5-HT and 2-methyl 5-HT were at least 5 times less potent to inhibit [ 3 H]GR67330 binding in rabbit ileum than rat brain. The most pronounced differences were evident with phenylbiguanide and m CPBG which were ∼ 70 and ∼ 300 times less potent in the rabbit ileum respectively compared with the rat tissues. These differences were unlikely to be due to depletion effects because tissue combination experiments (rabbit ileum and rat brain) yielded biphasic inhibition curves for phenylbiguanide with affinities for each component similar to those in the individual tissues. Antagonist affinities also varied between the rabbit and rat tissues, although less markedly. Amongst the antagonists, the most marked differences were apparent with SDZ 206–830 and quipazine each being ∼ 10 times less potent to inhibit binding to rabbit than rat tissue. Hill coefficients for inhibition of binding varied with tissue. In rat brain, as previously described for [ 3 H]GR67330, Hill coefficients for agonist (and quipazine) inhibition of binding were greater than unity. This was less marked in rat and rabbit ileum tissues. The present studies provide further evidence for species variation in 5-HT 3 receptors.

Profiles of interaction of R(+)/S(-)-zacopride and anxiolytic agents in a mouse model

The mouse black and white test box was used to measure changes in behaviour in an aversive situation where the administration of R(+)-zacopride (but not S(-)-zacopride) alone decreased aversive responding to the white area. A similar anxiolytic profile of action was observed using parachlorophenylalanine (PCPA), whose effects were antagonised by a co-treatment with R(+)-zacopride and reversed by S(-)-zacopride to an exacerbation of the aversive response. An anxiolytic profile of action was also observed using ondansetron, granisetron, chlordiazepoxide, diazepam, ritanserin, 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin), E4424 (2-[4-[4-(4-chloro-l-pyrazoyl)butyl]-l-piperazinyl]-pyrimidine), umepsirone, DuP753 (2-n-butyl-4-chloro-5-hydroxy-methyl-1-[2(1H-tetrazol-5-yl) biphenyl-4-yl)methyl)]-imidazole), SQ29,852 ((S)-1-[6-amino-2[hydroxy)(4-phenyl-butyl)phosphinyl]-oxy)-1- nexy]-2-proline), devazepide and guanfacine, and this was retained following co-treatment with PCPA. The anxiolytic profile of action of PCPA was also retained following co-treatment with renzapride which when administered alone failed to modify behaviour. However, the ability of chlordiazepoxide, diazepam, ondansetron and E4424 (but not devazepide, DuP753 or SQ29,852) to reduce aversive responding was inhibited by co-treatment with R(+) and/or S(-)-zacopride. It is concluded that the reduction in aversive responding caused by pharmacological manipulation at the benzodiazepine, 5-HT receptor subtypes 5-HT1A, 5-HT1C/5-HT2 and 5-HT3 (but not at the cholecystokin CCKA or angiotensin receptors or inhibition of angiotensin converting enzyme) can be inhibited by R(+) and S(-)-zacopride. The data is discussed in terms of zacopride having an agonist or partial agonist effect at the 5-HT3 receptor.

The profiles of interaction of yohimbine with anxiolytic and putative anxiolytic agents to modify 5-HT release in the frontal cortex of freely-moving rats.

1 The interaction of yohimbine with anxiolytic and putative anxiolytic agents to modify 5‐hydroxytryptamine (5‐HT) release in the frontal cortex of the freely‐moving rat was assessed using the microdialysis technique. 2 The α2‐adrenoceptor antagonist, yohimbine (5.0 mg kg−1, i.p.) increased maximally the extracellular levels of 5‐HT in the rat frontal cortex by approximately 230% of the basal levels. 3 The α2‐adrenoceptor agonist, clonidine (30–100 μg kg−1, i.p.) decreased dose‐dependently the extracellular levels of 5‐HT in the rat frontal cortex by approximately 0–60% of the basal levels. A 5 min pretreatment with clonidine (50 μg kg−1, i.p.) prevented the yohimbine‐induced increase in the extracellular 5‐HT levels. 4 The benzodiazepine receptor agonist, diazepam (2.5 mg kg−1, i.p.) and the 5‐HT3 receptor antagonist, ondansetron (100 μg kg−1, i.p.) (5 min pretreatment) completely prevented the yohimbine (5.0 mg kg−1, i.p.)‐induced increases in the extracellular levels of 5‐HT. The 5‐HT1A receptor agonist, 8‐OH‐DPAT (0.32 mg kg−1, s.c.) partially antagonized the yohimbine response. 5 A 5 min pretreatment with the 5‐HT3/5‐HT4 receptor ligand R(+)‐zacopride (10 μg kg−1, i.p.) reversed the yohimbine (5.0 mg kg−1, i.p.)‐induced increase in the extracellular levels of 5‐HT to approximately 30% below the basal levels. A 5 min pretreatment with S(−)‐zacopride (100 μg kg−1, i.p.) failed to modify the response to yohimbine. 6 The present study provides evidence of the ability of the anxiogenic agent, yohimbine, to increase the activity of the central 5‐hydroxytryptaminergic system and the ability of clonidine and various anxiolytic and putative anxiolytic agents to prevent the yohimbine response.

Actions of 5-hydroxytryptophan to inhibit and disinhibit mouse behaviour in the light/dark test

The involvement of 5-HT receptors in behavioural responding to an aversive situation was investigated in the mouse light/dark test. The administration of 5-hydroxytryptophan (5-HTP) (12.5-50 mg/kg i.p.) increased brain 5-HT turnover and inhibited mouse behaviour in the light/dark test box. The 5-HT2C/5-HT2A receptor antagonists methysergide (1.0 and 5.0 mg/kg i.p.) and ritanserin (0.1-1.0 mg/kg i.p.) antagonised (methysergide) or reversed (ritanserin) the effects of 5-HTP to an increased exploration of the light compartment; a low dose of the 5-HT3 receptor antagonist ondansetron (0.01 mg/kg i.p.) had a similar effect. The disinhibitory effect of the 5-HTP/ritanserin interaction was antagonised by the 5-HT3/5-HT4 receptor antagonists SDZ205-557 (0.001-0.1 mg/kg) and a high dose of tropisetron (1.0 mg/kg i.p.) but not by ondansetron (1.0 mg/kg i.p.). At these doses tropisetron and ondansetron had no effect in their own right. Thus the dominant effect of 5-HTP in the mouse is to inhibit behaviour, a response mediated via 5-HT2C/5-HT2A and 5-HT3 receptors. A 5-HT4 receptor may effect an opposing disinhibitory potential as revealed by ritanserin.

Toxic effects of solstitialin a 13-acetate and cynaropicrin from Centaurea solstitialis L. (asteraceae) in cell cultures of foetal rat brain

Extracts of the aerial parts of the yellow star thistle (Centaurea solstitialis L.) were prepared using petroleum ether, chloroform and methanol and toxicity assessed in primary cell cultures of striatum, frontal cortex and mesencephalon, containing the substantia nigra or raphe nucleus from the brain of the foetal rat. Chloroform extracts significantly reduced the percentage of live cells whereas the petroleum ether extract was inactive. The ability of the methanol extract to reduce the percentage of live cells could not be distinguished from the effects of the vehicle controls for methanol. Subsequently, the plant material was extracted with dichloromethane and 10 fractions were obtained by column chromatography. Fractions 5, 6, 7 and 8 caused concentration-dependent cell death in the culture of substantia nigra, the other fractions were not toxic at the concentrations used. In further studies, solstitialin A was isolated from fraction 9, solstitialin A 13-acetate from fraction 6 and solstitialin A-3 acetate and cynaropicrin from fraction 7. Solstitialin A 13-acetate and cynaropicrin were toxic to cultures of substantia nigra cells. It is concluded that, of the compounds identified, solstitialin A 13-acetate and cynaropicrin have toxic potential in cell cultures, containing cells from the substantia nigra of the rat, the specificity of action to cells of the substantia nigra remains to be shown, and that a toxic action in the midbrain may contribute to the nigro-pallidal encephalomalacia, caused by the ingestion of the yellow star thistle by horses.

The effect of 5-HT receptor ligands on the uptake of [3H]5-hydroxytryptamine into rat cortical synaptosomes

The effect of 5-HT receptor agonists and antagonists to inhibit [3H]5-HT uptake was investigated in rat cortical synaptosomes. The 5-HT (5-hydroxytryptamine) uptake inhibitors paroxetine and fluoxetine yielded pKi values of 8.41 +/- 0.12 and 7.43 +/- 0.06 respectively. The 5-HT3/5-HT4 receptor antagonist tropisetron and the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) had similar inhibitory potencies to cocaine (pKi values of 6.58 +/- 0.04, 6.47 +/- 0.14 and 6.45 +/- 0.12 respectively). The dopamine and noradrenaline uptake inhibitors GBR12909 and desipramine had comparable values of 6.5 +/- 0.05 and 6.13 +/- 0.07. Other 5-HT receptor ligands had pKi values less than 6.0 (R(+)-zacopride, MDL72222, R(+)/S(-)-zacopride) or 5.0 (5-methoxytryptamine, m-chlorophenylbiguanide, S(-)-zacopride, SDZ205-557, ondansetron and renzapride). It is concluded, with the possible exception of tropisetron and 8-OH-DPAT, that it is unlikely that the effects of the 5-HT receptor ligands to inhibit 5-HT uptake contribute to their effects in vivo.