David A.S Parker

Comparative activities of the enantiomeric GABAB receptor agonists CGP 44532 and 44533 in central and peripheral tissues

In neocortical slices maintained in Mg(2+)-free Krebs medium, the gamma-aminobutyric acid (GABA(B)) receptor agonists baclofen, (3-amino-2(S)-hydroxypropyl)methylphosphinic acid (CGP 44532), and its (R)-enantiomer CGP 44533 depressed the frequency of spontaneous discharges in a concentration-dependent manner (EC(50)=10, 6.5, and 50 microM, respectively). These effects were reversibly antagonised by the GABA(B) receptor antagonist (+)-(S)-5,5 dimethylmorpholinyl-2-acetic acid (Sch 50911) (3, 10, and 30 microM) (average pA(2) value=6.0+/-0.2). In neocortical wedges, baclofen, CGP 44532 and CGP 44533 elicited concentration-dependent hyperpolarisations (the EC(50)s were 14, 7.5 and 16 microM, respectively) sensitive to Sch 50911 (1, 5, 10 microM) (average pA(2) value=6.0+/-0.1), whilst they also depressed ileal electrically elicited cholinergic twitch contractions (EC(50)=11, 7, and 50 microM) that were antagonised by Sch 50911 (average pA(2) value=6.0+/-0.1). In electrically stimulated brain slices preloaded with [3H]GABA, baclofen, CGP 44532 and CGP 44533 decreased [3H]GABA release (IC(50)=5, 0.45, and 10 microM); this effect was reversed by Sch 50911 (50 microM). It is concluded that CGP 44532 is a far more potent agonist at GABA(B) autoreceptors than at central or peripheral heteroreceptors.

The morpholino-acetic acid analogue Sch 50911 is a selective GABAB receptor antagonist in rat neocortical slices

The pharmacological properties of (+)-(S)-5,5-dimethylmorpholinyl-2-acetic acid (Sch 50911) were evaluated on GABA(B) receptors in rat neocortical slices. The GABA(B) receptor agonist, baclofen, produced a concentration-dependent depression of the frequency of spontaneous discharges in slices maintained in Mg2+-free Krebs medium with an EC50 of 6 microM, reversibly antagonised by Sch 50911 (5, 10 and 25 microM) with an apparent pA2 of 6.0 +/- 0.1. The (-) enantiomer Sch 50910 (500 microM) and the racemic des-methyl analogue Sch 48588 (500 microM) were inactive. In slices preloaded with [3H]GABA, Sch 50911 antagonised GABA(B) autoreceptors, increasing the electrically-stimulated 3H overflow in a concentration-dependent manner, with an IC50 of 3 microM. The maximal effect (148 +/- 10.5%) was found at 10 microM, but at 50 microM the response was reduced to 67 +/- 19%. In contrast, evoked release was unaffected by Sch 50910 (100 microM) whilst Sch 48588 at 100 microM increased the overflow by 51.3 +/- 11.6%. In summary, Sch 50911 is a relatively potent antagonist of considerable potential in studies of GABA(B) receptor function.

CGP 36216 is a selective antagonist at GABAB presynaptic receptors in rat brain

In rat neocortical preparations maintained in Mg(2+)-free Krebs medium, baclofen depressed the frequency of spontaneous discharges in a concentration-dependent manner (EC(50) = 6 microM), sensitive to (3-aminopropyl)ethylphosphinic acid (CGP 36216) (100, 300 and 500 microM) (pA(2) = 3.9 +/- 0.1). By contrast, CGP 36216, up to 1 mM, was ineffective in antagonising baclofen-induced hyperpolarisations, mediated through gamma-aminobutyric acid(B) (GABA(B)) postsynaptic receptors. In electrically stimulated brain slices preloaded with [3H]GABA, CGP 36216 increased [3H]GABA release (IC(50) = 43 microM), which was reversed by baclofen (20 microM). While CGP 36216 is ineffective at GABA(B) postsynaptic receptors, it is appreciably more active at presynaptic receptors.

The CGP7930 analogue 2,6-di-tert-butyl-4-(3-hydroxy-2-spiropentylpropyl)-phenol (BSPP) potentiates baclofen action at gabab autoreceptors

1 The pharmacological actions of 2,6‐di‐tert‐butyl‐4‐(3‐hydroxy‐2‐spiropentylpropyl)‐phenol (BSPP), a putative presynaptic GABAB receptor modulator, were examined in electrically stimulated rat neocortical brain slices preloaded with [3H]‐GABA or [3H]‐glutamic acid. 2 At 10 mmol/L, BSPP inhibited the release of [3H]‐GABA in the presence of baclofen, but not that of [3H]‐glutamic acid. This effect was sensitive to the GABAB receptor antagonist (+)‐(S)‐5,5‐dimethylmorpholinyl‐2‐acetic acid (Sch 50911). 3 Alone, BSPP had no effect on the release of [3H]‐GABA or [3H]‐glutamic acid. 4 It is concluded that BSPP selectively potentiates the action of baclofen at GABAB autoreceptors, but not heteroreceptors and may be a useful ligand to discriminate between presynaptic GABAB receptor subtypes.

Cocaine inhibits extraneuronal O-methylation of exogenous norepinephrine in nasal and oral tissues of the rabbit.

Nasal mucosa (respiratory and olfactory) and lingual gingiva of the rabbit were depleted of their sympathetic nerves by superior cervical ganglionectomy. In the innervated nasal mucosa, exogenous tritiated norepinephrine (3H-NE) was metabolised mainly to tritiated 3,4-dihydroxyphenylethylene glycol (3HDOPEG) and 3,4-dihydroxy mandelic acid (3HDOMA), whereas after denervation it was metabolised mainly to tritiated normetanephrine (3HNMN). In the denervated mucosa, cocaine (30 umol/1) inhibited 3HNMN formation by 50-60%. Cocaine also inhibited 3HNMN formation by 60% in the denervated lingual gingiva. It is concluded that the tissues metabolise 3H-NE via a cocaine-sensitive extraneuronal uptake and O-methylating system similar to that which has been shown to be present in dental pulp.

Antagonism of GABAB receptors by morpholino-2-acetic acid derivatives Sch 54679 and Sch 51324 in rat brain

In rat neocortical slices maintained in Mg2+-free Krebs medium, baclofen depressed the rate of spontaneous discharges in a concentration-dependent manner (EC50 = 4.5 microM). This depression was reversibly antagonised by 5-(S,R)-hydroxymethyl-5-methylmorpholinyl-2-(R,S)-acetic acid (Sch 54679) and 2-(R,S)-5-[spirocyclopentyl]-morpholinyl-acetic acid (Sch 51324) (respective pA2 values of 5.8+/-0.15 and 5.4+/-0.2). In electrically-stimulated slices preloaded with [3H]gamma-aminobutyric acid (GABA), Sch 54679 (EC50 = 3 microM) was 2.3 times more potent than Sch 51324 (EC50 = 7 microM) in increasing [3H]GABA release through antagonism of GABA(B) autoreceptors. These structurally novel analogues may be pharmacologically useful for elucidating GABA(B) receptor functions.

Pharmacological actions of thymol and an analogue at GABAB autoreceptors

GABAB autoreceptors inhibit release of GABA from GABAergic nerve terminals. Agonists of these receptors (e.g. baclofen) inhibit, whereas antagonists (e.g. (+)‐(S)‐5,5‐dimethylmorpholinyl‐2‐acetic acid; Sch 50911) enhance release of the transmitter. The actions of thymol (2‐isopropyl‐5‐methylphenol) and the structurally related compound 2‐tert‐butyl‐4‐methylphenol, (4MP) on the release of [3H]‐GABA were examined in rat neocortical slices where the GABAergic nerves had been preloaded with [3H]‐GABA and subsequently stimulated electrically on two occasions (S1 and S2). Test agents, baclofen and Sch 50911 were added to the superfusion medium prior to the second period of stimulation (S2). Stimulation‐induced overflow (SIO) of [3H]‐GABA as a consequence of these stimulations (SIO1 and SIO2) were calculated and the effects of agents determined by comparing the SIO2/SIO1 ratio in the presence of each agent with that in control tissue. Thymol potentiated the release of [3H]‐GABA (EC50 170 μmol/L), an action reversed by baclofen (2 μmol/L). Baclofen alone had little effect on GABA release. Release of [3H]‐GABA was inhibited by 4MP (IC50 3 μmol/L) and this effect was blocked by Sch 50911 (10 μmol/L). Alone, Sch 50911 markedly potentiated the release of GABA. These results imply that 4MP is an agonist of GABAB autoreceptors; however, further studies are needed to confirm that thymol is indeed a GABAB autoreceptor antagonist. Of interest are structural differences in these agents. Thymol has a propyl group in the ortho position relative to the phenolic hydroxyl, whereas in 4MP this is a butyl group and the methyl group moves from position 5 to 4. Whether one or both of these changes was responsible for the above actions is unknown.

Pharmacological re-evaluation of a GABAB receptor antagonist CGP 47332A in rat brain

In rat neocortical slices maintained in Mg(2+)-free Krebs medium, the gamma-aminobutyric acid (GABA(B)) receptor agonist baclofen concentration-dependently depressed the frequency of spontaneous discharges (EC(50)=12 microM). This was reversibly antagonised by (R, S)-3-amino-2-hydroxy-propyl-P-n-butyl-phosphinic acid (CGP 47332A) (25, 100, 300 microM) which produced rightwards shifts of the baclofen concentration-response curves (pA(2) value=4.8+/-0.1). In electrically stimulated slices preloaded with [3H]GABA, CGP 47332A increased its release (EC(150)=100 microM) through antagonism of GABA(B) autoreceptors. Although CGP 47332A was some six times weaker on GABA(B) auto- than on heteroreceptors, yet its congener lacking the beta-hydroxy substituent displays equal potency in both binding (IC(50)=38 microM) and GABA(B) autoreceptor functional studies (EC(150)=38 microM) as previously reported [Froestl, W., Mickel, S.J. , Von Sprecher, G., Diel, P.J., Hall, R.G., Maier, L., Strub, D., Melillo, V., Baumann, P.A., Bernasconi, R., Gentsch, C., Hauser, K., Jaekel, J., Karlsson, G., Klebs, K., Maitre, L., Marescaux, C., Pozza, M.F., Schmutz, M., Steinmann, M.W., Van Riezen, H., Vassout, A., Mondadori, C., Olpe, H.R., Waldmeier, P.C., Bittiger, H., Phosphinic acid analogues of GABA: 2. Selective, orally active GABA(B) antagonists. J. Med. Chem. 38 (1995) 3313-3331.].

Inhibition by cocaine of noradrenaline release from sympathetic nerves in the rabbit ear artery.

SUMMARY

Pharmacological actions of oximino-propofol analogues at GABAB autoreceptors

1. GABAB autoreceptors are a subclass of GABAB receptors that inhibit the release of [3H]GABA from GABAergic nerve terminals. Baclofen is an agonist that reduces [3H]GABA, whilst the antagonist (+)‐(S)‐5,5‐dimethylmorpholinyl‐2‐acetic acid (Sch 50911) enhances [3H]GABA release in electrically‐stimulated rat neocortical brain slices preloaded with [3H]GABA. Here, the pharmacological actions of a series of compounds derived from the positive allosteric modulator, 2,6‐di‐tert‐butyl‐4‐(3‐hydroxy‐2,2‐dimethyl‐propyl)‐phenol (CGP7930), were examined on GABAB autoreceptors.