David J. Barber

Dexamethasone induces limited apoptosis and extensive sublethal damage to specific subregions of the striatum and hippocampus: implications for mood disorders.

It has been shown previously that the synthetic corticosteroid dexamethasone induces apoptosis of granule cells in the dentate gyrus and striatopallidal neurons in the dorsomedial caudate-putamen. We investigated whether or not dexamethasone can induce damage to other neuronal populations. This issue was addressed using OX42 immunohistochemistry to visualise activated microglia and thereby gauge the extent of dexamethasone-induced neuronal death. A single dose of dexamethasone (20mg/kg, i.p.) administered to young male Sprague-Dawley rats induced a strong microglial reaction which was restricted to the striatum, the dentate gyrus and all of the CA subfields of the hippocampus. Some OX42-immunoreactive cells were also seen in the lateral septal nucleus. Subsequent quantitative analysis of silver/methenamine-stained sections confirmed that acute administration of dexamethasone induced apoptosis in the striatum and all regions of the hippocampus at doses as low as 0.7mg/kg. In contrast, dexamethasone failed to induce apoptosis in the lateral septal nucleus at doses up to 20mg/kg. The levels of dexamethasone-induced striatal and hippocampal apoptosis were attenuated by pretreatment with the corticosteroid receptor antagonist RU38486 (Mifepristone), which implies that the cell death was mediated by a corticosteroid receptor-dependent process. We further determined whether dexamethasone induced sublethal damage to neurons by quantifying reductions in the number of microtubule-associated protein-2-immunoreactive striatal and hippocampal cells following injection of the corticosteroid. Dexamethasone induced dramatic decreases in the striatum, with the dorsomedial caudate-putamen being particularly affected. Similar damage was seen in the hippocampus, with the dentate gyrus and CA1 and CA3 subfields being particularly vulnerable.Equivalent corticosteroid-induced neuronal damage may occur in mood disorders, where the levels of endogenous corticosteroids are often raised. Corticosteroid-induced damage of striatal and hippocampal neurons may also account for some of the cognitive deficits seen following administration of the drugs to healthy volunteers.

Benzodiazepine receptor ligands and the consumption of a highly palatable diet in non-deprived male rats

Non-deprived rats were familiarised with a highly palatable diet until baseline consumption in a 60-min daily access period had stabilised. The benzodiazepine receptor agonist midazolam (1.25–10.0 mg/kg, IP) produced a large, dose-related increase in food consumption during the first 30 min of access. It also produced significant, short-term hyperphagia in animals which had been partially pre-satiated on the diet before drug administration, an effect which was reversible by the benzodiazepine receptor antagonist Ro15-1788. Administered alone, Ro15-1788 (1.25–10.0 mg/kg, IP) had no intrinsic activity in the food consumption test. In contrast, CGS 8216 (2.5–40.0 mg/kg, IP) produced a marked dose-related suppression of food intake. This anorectic effect was shared by two benzodiazepine receptor inverse agonists, FG 7142 and DMCM, which also produced dose-dependent reductions in consumption. The effects on feeding produced by FG 7142 (20 mg/kg, IP) and DMCM (1.25 mg/kg, IP) were reversed by either Ro15-1788 (2.5 and 5.0 mg/kg) or midazolam (5.0 and 10.0 mg/kg). A matched anorectic effect produced by CGS 8216 (40 mg/kg) was not, however, reversed by either Ro15-1788 or midazolam. This suggests that at a high dose CGS 8216 may act by a mechanism different from that of the two inverse agonists. The feeding test described in the report proved sensitive to both hyperphagic and anorectic effects of drugs active at benzodiazepine receptors, pointing to a possible bi-directional control of palatable food consumption.

Reversal of the anorectic effect of (+)‐fenfluramine in the rat by the selective cholecystokinin receptor antagonist MK‐329

1 Experiments were conducted to determine whether or not the effect of (+)‐fenfluramine (3.0 mg kg−1, i.p.) on food intake can be antagonized by the selective cholecystokinin receptor antagonist MK‐239 (formerly L364/718; (3S(−)‐N‐(2,3‐dihydro‐1‐methyl‐2‐oxo‐5‐phenyl‐1‐H‐1,4‐benzodiazepin‐3‐yl)‐1H‐indole‐2‐carboxamide). Two feeding paradigms were employed. In the first, non‐deprived rats were familiarized with eating a highly palatable, sweetened mash in a 30 min test. In the second, freely‐feeding rats were trained to consume powdered chow in their home‐cages, and their intake was monitored over the first 6 h of the night‐period. 2 In doses of 30.0 and 100.0 μg kg−1, s.c., MK‐329 almost completely blocked the anorectic effect of (+)‐fenfluramine in the palatable food intake test. These doses of MK‐329 have previously been reported to antagonize the anorectic effect produced by exogenous cholecystokinin‐octapeptide (CCK8) in rats. Both doses of MK‐329 were also effective in significantly attenuating the anorectic effect of (+)‐fenfluramine in nocturnal free‐feeding animals over a 6 h‐period. 3 MK‐329 (10.0–100.0 μg kg−1, s.c.) failed to antagonize the anorectic effect of either the specific dopamine D2‐receptor agonist quinpirole (0.3 mg kg−1, s.c.) or the β‐carboline FG 7142 (10.0 mg kg−1, i.p.) in the palatable food intake test. 4 MK‐329 (10.0–300.0 μg kg−1, s.c.) had no effect, when administered alone, on the level of palatable food intake in non‐deprived rats, even when substantial satiation was produced by a pre‐feeding procedure. Furthermore, MK‐329 had no effect, when administered alone, on nocturnal food intake in freely‐feeding rats. 5 In conclusion, not only was MK‐329 a potent antagonist of the effect of CCK8 on food intake, it also blocked the effect of (+)‐fenfluramine to a significant degree. The effect of MK‐329 was selective in that the anorectic effects of either quinpirole or FG 7142 remained unaffected. Administered alone, MK‐329 did not affect food intake, indicating that its reversal of (+)‐fenfluramine‐induced anorexia was not secondary to an intrinsic hyperphagic effect. The results provide some evidence that the depressant effect of (+)‐fenfluramine on food intake depends on the activity of endogenous CCK.

Asymmetric generalization between the discriminative stimulus effects of nicotine and cocaine.

The discriminative stimulus effects of nicotine and cocaine were studied, alone and in combination, in rats. Two sets of rats were trained to press one lever when injected intraperitoneally (i.p.) with either nicotine (0.1 mg/kg = 0.6 micromol/kg, Set 1) or cocaine (8.9 mg/kg base = 29.4 micromol/kg, Set 2), and another lever when injected with saline. Rats learned to discriminate drug from saline, and maintained discriminative control throughout the study (at > 85% drug-appropriate responding). In accordance with most previous findings, cocaine only partially substituted for nicotine (maximum = 41% nicotine-lever responding). The nicotinic agonist, nornicotine, produced dose-related, near-full substitution for nicotine (maximum = 76% nicotine-lever responding), whereas the peripherally acting nicotinic agonist, methylcarbamylcholine, did not substitute for nicotine. The muscarinic receptor agonist pilocarpine also failed to substitute for nicotine. However, in the cocaine-trained rats, nicotine substituted fully for cocaine in a dose-dependent manner, demonstrating that cross-generalization between the two drugs is not symmetrical. Finally, administration of each drug as a pre-treatment to the other yielded inconsistent increases in each drugs discriminative stimulus effects. The results are congruent with the view that the discriminative stimulus effects of nicotine and cocaine share common features, but the asymmetric pattern of cross-generalization and the interactions revealed in the combination tests also suggest that there are important differences between them.

17β-oestradiol attenuates dexamethasone-induced lethal and sublethal neuronal damage in the striatum and hippocampus

Abnormal corticosteroid release is extensively associated with mood disorders. This association may result from the toxic actions of endogenous corticosteroids which can induce apoptosis of hippocampal neurons. Similarly, dexamethasone, a synthetic corticosteroid, can induce lethal and sublethal damage to rat hippocampal and striatal neurons and can result in steroid-induced psychoses in humans. The experiments reported here tested the hypothesis that pre-treatment with oestrogen would also attenuate dexamethasone-induced neuronal damage as oestrogens have neuroprotective actions against a variety of insults and falling levels of oestrogen are associated with increased vulnerability to mood disorders. Male Sprague-Dawley rats received three systemic injections which were a combination of vehicle, 17-beta-oestradiol (0.2 mg/kg, s.c.), the oestrogen receptor antagonist tamoxifen (10 mg/kg, s.c.) and dexamethasone (0.7 mg/kg, i.p.) and were killed 24 h after the final injection. Injections of dexamethasone (when preceded by vehicle injections) resulted in elevated levels of apoptosis and sub-lethal damage, as demonstrated by reduced levels of microtubule-associated protein-2-immunopositive neurons, in the striatum and hippocampus. This damage was regional with the dorsomedial caudate putamen and the dentate gyrus and CA1 and CA3 hippocampal sub-fields being particularly affected. Pretreatment with oestrogen substantially attenuated the dexamethasone-induced neuronal damage. This oestrogen-induced neuronal protection was in turn virtually eliminated by giving an initial injection of tamoxifen. These results suggest, therefore, that oestrogens can protect from corticosteroid-induced neuronal damage via an oestrogen receptor-mediated process.

Effects of baclofen on feeding behaviour examined in the runway

The motivational mechanisms underlying the effects of systemic administration of the GABA-B agonist baclofen on feeding were examined using a runway. Food-deprived male hooded Lister rats were trained to traverse a runway for food reinforcement. Baclofen (1 mg/kg i.p.) significantly increased food intake and this was most evident on the final two blocks of testing. The 2 mg/kg dose of baclofen increased running speed without significantly altering intake. At the highest dose tested (4 mg/kg), no significant effects on either consummatory or appetitive measures were observed. These data suggest that low doses of baclofen enhance the consummatory phase of ingestion by attenuating the natural signals associated with onset of satiation. The data also suggest that baclofen has complex effects on appetitive behaviour that may interfere with its effects on consumption.

Yawning induced by the selective dopamine D2 agonist N-0437 is blocked by the selective dopamine autoreceptor antagonist (+)-UH 232

Yawning and stretching responses were elicited in rats by a small dose (0.3 mg/kg) of the highly selective dopamine D2 agonist, N-0437. The responses were blocked by the highly selective dopamine autoreceptor antagonist, (+)-UH 232 (3.0 mg/kg), but not by raclopride at a dose which selectively blocks postsynaptic D2 receptors. The results strongly confirm the view that yawning and stretching are behavioral responses elicited by stimulation of presynaptic D2 receptors.

Differential effects of two cannabinoid receptor agonists on progressive ratio responding for food and free-feeding in rats

The cannabinoid receptor agonists Δ9-tetrahydrocannabinol (Δ9-THC) and HU-210 were compared in terms of their effects on: (1) progressive ratio (PR) responding for food, and (2) free food intake. In the first experiment, food-deprived Wistar rats were trained on a time-constrained (60 min) PR-5 schedule for food reinforcement, in which the response requirement incremented by five lever presses for each successive reinforcer. One group of rats received vehicle, 0.5, 1 or 3 mg/kg Δ9-THC (i.p.), and three other groups received HU-210 (i.p.) at three different dose ranges, spanning 0.001−0.1 mg/kg. In the second experiment, the effects of the two drugs on free food intake were tested in a separate group of non-deprived rats. For PR responding, Δ9-THC significantly increased the break point (final ratio completed) and the total number of lever presses emitted. The same drug also significantly increased free food intake. However, the effects of HU-210 were quite different: it did not alter PR responding at any dose; instead, its only significant effect was to reduce free food intake at 0.06 mg/kg. These data suggest that increased motivation to obtain food might underlie the hyperphagic effects of Δ9-THC. However, the synthetic agonist HU-210 has different effects: it only acts to reduce feeding behaviour, an outcome that probably reflects non-specific behavioural disruption. These findings suggest important differences between the two CB1 receptor agonists in terms of their pharmacological effects.

Effects of dl-fenfluramine and xylamidine on gastric emptying of maintenance diet in freely feeding rats

Freely feeding rats received an anorexigenic dose of dl-fenfluramine HCl (5 mg/kg). Two hours following injection, their stomachs retained significantly greater dry weight contents than saline-injected controls. The same dose of fenfluramine decreased the rate of gastric emptying over a 2 h period to a similar extent in mildly food-deprived rats. The peripherally acting serotonin antagonist xylamidine counteracted the effect of fenfluramine in prolonging the satiating effect of an ad libitum meal of a given size. We propose therefore that the principal mechanism by which fenfluramine reduces food consumption in freely feeding rats is through a prolongation of the satiating effect of absorption as a result of slowing of gastric emptying, presumably via enhanced release of serotonin from nerve terminals in the wall of the gastrointestinal tract.

Fluoxetine reduces food intake by a cholecystokinin-independent mechanism

The selective serotonin uptake inhibitor, fluoxetine (3.0-10 mg/kg), produced a significant dose-related suppression of palatable food consumption in nondeprived rats. The anorectic effect of fluoxetine (10 mg/kg) was not reversed by the potent and highly selective cholecystokinin receptor antagonist MK-329 [1-methyl-3-(2-indolyl) amino-5-phenyl-3H-1,4-benzodiazepin-2-one], administered in doses of 10-100 micrograms/kg. Fluoxetine (10 mg/kg) also significantly reduced the consumption of powdered laboratory chow in a 6-hr nocturnal free-feeding test. The anorectic effect in this paradigm was also not antagonized by MK-329. In contrast to previous data for d-fenfluramine (which enhances serotonin release), these results indicate that fluoxetine may suppress food intake by a mechanism which is independent of endogenous cholecystokinin.