Characterization of 5‐HT3 and ‘atypical’ 5‐HT receptors mediating guinea‐pig ileal contractions in vitro
Authors: Eglen R.M. , Swank S.R. , Walsh L.K.M. , Whiting R.L. 
Published: July 2012

Neuronal 5‐hydroxytryptamine (5‐HT) receptors mediating contraction of guinea‐pig ileal segments have been characterized in vitro by the use of methysergide to block 5‐HT1‐like and 5‐HT2 receptors. Concentration‐response curves to 5‐HT were biphasic (first phase, defined as those responses occurring between 1 nm and 0.32 μm 5‐HT, –log EC50 = 7.15 ± 0.08; second phase, defined as these responses occurring between 0.32 μm and 32 μm 5‐HT, –log EC50 = 5.32 ± 0.03) but monophasic to 5‐methoxytryptamine (−log EC50 = 7.0 ± 0.08) and 2 methyl 5‐HT (−log EC50 = 5.2 ± 0.13). The maximal response of the first phase to 5‐HT and the maximal response to 5‐methoxytryptamine were 30 ± 4% and 35 ± 5% respectively of the maximum response to the second phase of the 5‐HT concentration‐effect curve (set at 100%). In contrast, the maximal response to 2‐methyl‐5‐HT equalled that obtained with 5‐HT (second phase).

The responses comprising the second phase of the concentration‐effect curve to 5‐HT were antagonized by 1 μm ICS 205–930, ondansetron, granisetron, quipazine, N‐methyl‐quipazine and (R,S)‐zacopride and the following pKB values, with 5‐HT as the agonist, were obtained at the 5‐HT3 receptor: ICS 205–930 7.61 ± 0.05, ondansetron 6.90 ± 0.04, granisetron 7.90 ± 0.04, (S)‐zacopride 8.11 ± 0.06, (R,S)‐zacopride 7.64 ± 0.11, and (R)‐zacopride 7.27 ± 0.06.

Under conditions of 5‐HT1‐like, 5‐HT2 and 5‐HT3 receptor blockade, the following rank order of agonism was observed: 5‐HT > 5‐methoxytryptamine = renzapride > (S)‐zacopride > (R,S)‐zacopride > 5‐carboxamidotryptamine > BRL 24682 > (R)‐zacopride > metoclopramide > 2‐methyl‐5‐HT ≫ sulpiride. 8‐Dihydroxydiphenylaminotetralin (8‐OHDPAT), GR 43175, N,N‐dipropyl‐5‐carboxamidotryptamine, ondansetron, ICS 205–930, granisetron, quipazine and N‐methyl‐quipazine were inactive as agonists and antagonists. Relative to 5‐HT, (R,S)‐zacopride acted as a partial agonist (intrinsic activity, α = 0.80; –log EC50 = 6.3 ± 0.12; –log KA = 6.1 ± 0.03) as did (R)‐zacopride (α = 0.4, –log EC50 5.7 ± 0.08, –log KA = 5.5 ± 0.11). (S)‐zacopride acted as a full agonist (−log EC50 = 6.9 ± 0.03). ICS 205–930 (3 μm) antagonized competitively responses to 5‐HT, 5 methoxytryptamine, (R,S)‐ and (S)‐zacopride and 5‐carboxamidotryptamine yielding –log KB estimates ranging from 6.1–6.5.

It is concluded that two different 5‐HT receptors mediate excitatory neuronal responses in the guinea‐pig ileum. 5‐HT3 receptors mediate the second phase of the biphasic concentration‐response curve, whereas a receptor with properties distinct from the 5‐HT1‐like, 5‐HT2 and 5‐HT3 subtypes mediates the initial phase of the concentration‐response curve. This receptor, which exhibits a close similarity to the 5‐HT4 subtype is: (1) stimulated by 5‐methoxytryptamine but not 2‐methyl‐5‐HT; (2) stimulated selectively by certain substituted benzamides; (3) recognizes the optical isomers of zacopride and (4) is blocked by relatively high concentrations ICS 205–930 (pKB = 6.0–6.5) but not ondansetron, granisetron, quipazine or N‐methyl‐quipazine.