Kevin Fone

The affinity of several antidepressant and antipsychotic drugs for the 5-HT7 receptor and its CNS distribution suggest potential in the treatment of psychiatric diseases. However, there is little direct evidence of receptor function in vivo to support this. We therefore evaluated 5-HT7 receptors as a potential drug target by generating and assessing a 5-HT7 receptor knockout mouse. No difference in assays sensitive to potential psychotic or anxiety states was observed between the 5-HT7 receptor knockout mice and wild type controls. However, in the Porsolt swim test, 5-HT7 receptor knockout mice showed a significant decrease in immobility compared to controls, a phenotype similar to antidepressant treated mice. Intriguingly, treatment of wild types with SB-258719, a selective 5-HT7 receptor antagonist, did not produce a significant decrease in immobility unless animals were tested in the dark (or active) cycle, rather than the light, adding to the body of evidence suggesting a circadian influence on receptor function. Extracellular recordings from hypothalamic slices showed that circadian rhythm phase shifts to 8-OH-DPAT are attenuated in the 5-HT7 receptor KO mice also indicating a role for the receptor in the regulation of circadian rhythms. These pharmacological and genetic knockout studies provide the first direct evidence that 5-HT7 receptor antagonists should be investigated for efficacy in the treatment of depression.

Intrathecal injection of the analogue of TRH, CG 3509, into conscious rats produced dose-related wet-dog shakes and forepaw licking, which showed a bell-shaped relationship of intensity to dose. Pretreatment with alpha-MPT intraperitoneally, markedly reduced levels of noradrenaline and dopamine in the spinal cord and brainstem and attenuated both CG 3509-induced responses, while intrathecal treatment with DSP4 selectively reduced noradrenaline in the spinal cord without affecting either behaviour. Since denervation supersensitivity may develop following treatment with DSP4, these results are not inconsistent with a proposal that bulbospinal noradrenergic neurones modulate the behaviour induced by CG 3509. Wet-dog shakes and forepaw licking induced by CG 3509 were reduced by pretreatment with phenoxybenzamine or prazosin, suggesting that a tonic noradrenergic pathway may facilitate both behavioural responses through alpha 1-adrenoceptors. Methoxamine, combined with CG 3509 partially attenuated the wet dog shake behaviour, but methoxamine produced marked hindlimb jerking which might physiologically antagonise wet-dog shakes. Concomitant administration of clonidine and CG 3509 potently reduced wet-dog shakes in a dose-related manner but did not significantly affect forepaw licking, while idazoxan did not significantly affect either response. The latter findings imply that alpha 2-adrenoceptors play different roles in modulating the two behavioural responses and the possible synaptic location of the receptors is discussed. Taken together these results suggest that CG 3509 may release noradrenaline from bulbospinal neurones regulating motor function.

Effects of repeated intracerebroventricular administration of the thyrotrophin-releasing hormone (TRH) analogue, RX77368 (3,3'-dimethyl-TRH, 2 μg, once daily), on a scopolamine-induced performance deficit in an eight-arm radial maze were evaluated in adult rats. Scopolamine (0.3 mg/kg i.p.—30 min) pre-treatment produced a significant deficit in the number of unrepeated arm entries and total arm entries and increased the percentage of incorrect arm entries and the total time on the maze, compared with saline-treated controls. Prior treatment with RX77368 (40 min before maze testing) produced a partial but significant attenuation of the scopolamine-induced performance deficit on the maze during the first five trials but RX77368 also enhanced maze performance during the same period when given alone. These results suggest that the observed scopolamine-induced performance deficit on the radial arm maze partly results from a reduction in locomotion and maze exploration rather than solely impairment of memory, and that RX77368 treatment may improve radial maze performance by increasing arousal and exploratory behaviour in rats rather than directly enhancing cognition.

Many studies point to an involvement of deficits in the serotonergic nervous system and hyperactivity of the hypothalamic‐pituitary‐adrenal (HPA) axis function with depression. Indeed early life stress, involving HPA axis activation, may predispose susceptible individuals to develop depression in later life. This study investigates the effects of elevating the neuroendocrine stress hormone, corticosterone, for 1 week in adolescent rats on markers of serotonergic neurone function at adulthood. Slow release corticosterone pellets were implanted for 7 days and various serotonergic parameters, as well as plasma corticosterone levels, were measured on day 7 or on day 28 (21 days following removal of the pellet). The corticosterone implant attenuated weight gain and reduced adrenal weights compared to that in control rats implanted with a cholesterol pellet. After 7 days, with the implant still in place, the diurnal variation in plasma corticosterone was reduced so that the level was approximately at that of the evening peak throughout the day. Twenty‐one days after removal of the implant, the diurnal variation in plasma corticosterone returned. Corticosterone treatment decreased [3H] 8‐hydroxy‐2‐(di‐n‐propylamino)tetralin binding to the 5‐hydroxytryptamine1A receptor in the cortex but not in the hippocampus. Corticosterone treatment also enhanced the circadian rhythm observed in 5‐hydroxyindoleacetic acid level and the ratio of 5‐hydroxyindoleacetic acid to the 5‐hydroxytryptamine in the frontal cortex. Despite corticosterone pellet removal 21 days earlier, there was a persistent decrease in whole body and adrenal weight, cortical 5‐hydroxytryptamine1A receptor binding and an alteration in the diurnal variation in the 5‐hydroxytryptamine ‘turnover’ in the frontal cortex.

This study examined the involvement of the 5-HT(2A) receptor in the long-term anxiogenic effect of a brief exposure of young rats to 3,4-methylenedioxymethamphetamine (MDMA) using the social interaction and elevated plus-maze paradigms. Wistar rats (post-natal day (PND) 28) received either MDMA (5 mg/kg i.p.) or saline (1 ml/kg i.p.) hourly for 4 h on 2 consecutive days. Locomotor activity was measured for 60 min after the first injection and core body temperature was recorded at regular intervals over 4 h. On PND 84, without further drug administration, social interaction was assessed between treatment-matched rat pairs derived from separate litters. On PND 86, rats received either the 5-HT(2A/2C) receptor agonist, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI, 1 mg/kg i.p.) or saline and locomotor activity, wet-dog shakes and back muscle contractions were monitored. The change in elevated plus-maze behaviour was assessed following the same injection on PND 87. Acutely, MDMA produced a significant hyperlocomotion and hyperthermia (p<0.01). Following 55 days of abstinence, social interaction was reduced by 27% in MDMA pre-treated rats compared with that in controls (p<0.01). On the elevated plus-maze, pre-treatment with MDMA prevented the anxiogenic effect of DOI. On PND 92, hippocampal, frontal cortical and striatal 5-hydroxytryptamine (5-HT) was significantly reduced in MDMA pre-treated rats by between 16% and 22%, without any accompanying change in [(3)H]paroxetine binding in cortical homogenates. In conclusion, exposure of young rats to repeated MDMA caused serotonin depletion and induced 'anxiety-like' behaviour in the social interaction test accompanied by a long-lasting reduction in specific 5-HT(2A) receptor mediated behaviour.

5-HT(6) receptors are expressed in brain regions associated with learning and memory, and blockade of their function increases central cholinergic and glutamatergic neurotransmission and enhances cognitive processes. This study examined the effects of acute systemic administration of two selective 5-HT(6) receptor antagonists Ro 04-6790 and SB-271046 (10 mg kg(-1) i.p.) on acquisition, consolidation, and retrieval in the novel object discrimination (NOD) task, a two-trial test of recognition memory in which rats exposed to two identical objects during a familiarisation trial can discriminate a novel from a familiar object during the subsequent choice trial, following inter-trial delays of up to 3 h. 5-HT(6) receptor antagonist administration 20 min prior to or immediately after the familiarisation trial, but not 20 min prior to the choice trial reversed the deficit in object discrimination produced by a 4 h inter-trial interval. The nootropic effects of the 5-HT(6) receptor antagonists in this task thus appear to involve enhanced consolidation. Pre-treatment with the non-competitive NMDA receptor antagonist MK-801 (0.05 mg kg(-1) i.p.) prevented the effect of Ro 04-6790 on delay-induced deficits in object discrimination. This suggests that the 5-HT(6) receptor antagonist-induced enhancement of consolidation involves increased central glutamatergic neurotransmission.

Hypercortisolism and altered serotonergic function may account for the pathological symptoms seen in depression. This study examines the impact of 4 days continuous corticosterone treatment on 5-HT agonist-induced behaviour to delineate changes in 5-HT receptor function in the adult rat. The flat body posture, reciprocal forepaw treading, elevated corticosterone, hyperglycaemia, hypothermia and reduced hippocampal 5-HT induced by the 5-HT(1A) agonist 8-OHDPAT (0.3 mg/kg ip) were all significantly attenuated by the corticosterone implant. The elevation in plasma corticosterone and back muscle contractions evoked by the 5-HT(2A) agonist DOI (1 mg/kg ip) were attenuated, whilst wet-dog shakes were enhanced by corticosterone treatment. 5-HT(2B) agonist-induced behaviour and the hypolocomotion and hypophagia induced by the 5-HT(2C) agonist m-CPP (2.5 mg/kg ip) were unaltered but the mCPP-induced elevation in corticosterone was abolished by corticosterone treatment. Hypothalamic 5-HT receptors mediating corticosterone- and 5-HT(1A) receptors, whether on serotonergic nerve terminals or postsynaptic neurones, were downregulated by corticosterone treatment. In contrast, 5-HT(2A) receptors may be up- or downregulated dependent on whether they are on supraspinal or spinal neurones, respectively. A comparison of the brain region-dependent alteration in serotonergic function produced by hypercorticosterone in the rat with that seen in depression is discussed.

The effect of repeated intrathecal injection of thyrotrophin-releasing hormone (TRH) and two analogues of TRH, C-terminally modified RX 77638 and N-terminally modified CG 3509, were examined on behavioural (wet-dog shakes and forepaw licking) and biochemical markers for spinal motoneurones, bulbospinal raphe nerve terminals and the pituitary-thyroid axis in rats. Saline (10 microliters washed in with 15 microliters), TRH (20 micrograms), RX 77368 (2 micrograms) or CG 3509 (2 micrograms) were administered intrathecally (twice daily for 3 or 5 days), after which levels of plasma-free thyroxine and thyroid-stimulating hormone (TSH) were measured and the dorsal and ventral portions of the thoracolumbar spinal cord, brainstem and hypothalamus were assayed for TRH- and calcitonin gene-related peptide (CGRP)-like immunoreactivity, levels of indoleamines and the activity of choline acetyltransferase (ChAT). Behavioural tolerance developed rapidly with consecutive injections of RX 77368, such that wet-dog shakes were significantly reduced and forepaw-licking tended to be decreased by the third intrathecal injection. Five, but not 3, days of administration of RX 77368 selectively elevated levels of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid in the ventral spinal cord, where these substances are principally located in bulbospinal raphe nerve terminals. The time course of the change in indoleamines suggests that administration of TRH peptides elevated the synthesis, rather than the release, of 5-HT from these nerve terminals.(ABSTRACT TRUNCATED AT 250 WORDS)

The 5-hydroxytryptamine (5HT) containing bulbospinal pathway was studied with immunohistochemical (IF) and chemical techniques in 2-3 and 30 months old male Sprague-Dawley rats. The coexisting neuropeptides substance P (SP), thyrotropin-releasing hormone (TRH) and galanin were also analysed. Furthermore, the expression of mRNA encoding aromatic L-amino acid decarboxylase (AADC), prepro-TRH, and preprotachykinin (prepro-SP) was analysed with in situ hybridization (ISH) in the midline raphé nuclei inthe lower brainstem. The results showed a decreased number of axonal 5HT fibers with a normal morphology in the ventral horn of the aged rat lumbosacral spinal cord, and several 5HT immunoreactive (IR) fibers with an aberrant morphology, suggestive of axonal degeneration, were intermingled. This was evident in both the dorsal and ventral horn of the spinal cord. The 5HT-IR fibers with an aberrant morphology usually also contained TRH-and/or SP- and/or galanin-like immunoreactivity (LI) in the ventral horn. These signs of degeneration were clearly less evident in the thoracic and cervical spinal cord segments. Moreover, these changes varied between aged litter-mates. This was in agreement with behavioural signs of motor disturbances, present in about 40% of the aged rats and which in all cases were confined to the hindlimbs. Chemical analyses disclosed significantly lower levels of TRH-LI and, in particular, SP-LI in both the ventral and dorsal quandrants of the spinal cord in the aged rat compared to young adults. The differences were largest in the lumbar regions of the spinal cord. Corresponding analysis of 5HT and 5-hydroxyindoleacetic acid (5HIAA) in the same tissue specimens revealed largely unaltered levels of 5HT and a slight increase in 5HIAA, indicating the possibility of an increased 5HT turnover in the aged rat spinal cord. Neurons in nucleus raphé obscurus and nucleus raphé pallidus were immunoreactive to 5HT, and after pretreatment with colchicine to TRH-, SP-, and galanin-LI as well. There was no obvious difference in number of labeled cells, or labeling intensity, between colchicine-treated young adult and aged rats, although, in the corresponding region of medulla oblongata, chemical analysis disclosed significantly lower levels of 5HT, TRH, and, in particular, SP in untreated aged rats. In contrast, in situ hybridization analysis revealed increased mRNA levels encoding prepro-TRH and prepro-SP in old rats, while mRNA content encoding AADC mRNA was similar in young adult and aged rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Postweaning social isolation in the rat induces lasting alterations that parallel several of the core symptoms seen in human schizophrenics, including hyperreactivity to novel environments, cognitive impairment, and deficits in sensorimotor gating. The current study determined whether these changes are accompanied by any elevation in the proportion of striatal dopamine receptors in the functional high affinity state (D(2) (High)), as observed in other preclinical models of psychosis. Male Lister hooded rats (20-24 days) were housed in groups of three or alone. On Day 36 postweaning locomotor activity was monitored for 60 min in a novel arena, and on Day 37 novel object discrimination was assessed using a 2 h intertrial interval. Three days later striata were collected, homogenized, washed three times to remove endogenous dopamine, and the proportion of D(2) (High) determined by competition between dopamine and 2.27 nM [(3)H]domperidone. Isolates were significantly more active than group housed controls for both ambulation and rears. Although both groups exhibited comparable levels of familiarization trial object exploration, group housed animals were able to discriminate between novel and familiar objects during the choice trial while isolates were not. Social isolation was associated with a highly significant elevation in the proportion of striatal D(2) (High), equivalent to a 3.3-fold increase (group 15.2% +/- 1.4%, isolate 49.8% +/- 4.8%; P < 0.0001, Student's unpaired t-test). These findings support both the hypothesis that elevated D(2) (High) is a common feature of multiple animal models of psychosis, and the validity of isolation rearing as a neurodevelopmental model of a "schizophrenia-like" state.

The tetrahydrobenzindole, 2a-(4-(4-phenyl-1,2,3,6-tetrahydropyridyl)butyl)-2a,3,4,5-tetrahydrobenzo[cd]indol-2(1H)-one (DR4004) has been described as a highly selective antagonist for the 5-hydroxytryptamine(7) (5-HT(7)) receptor [J. Med. Chem. 42 (1999) 533]. Consistent with original data, DR4004 bound to rat hypothalamic membranes with an affinity of 7.3+/-0.2 (pK(i)+/-S.E.M.) for the 5-HT(7) receptor. However, competition binding studies showed that DR4004 had poor receptor selectivity with the following affinity profile; dopamine D2 receptor, alpha(1)-adrenoceptor > or =5-HT(7) receptor>histamine H(1) receptor, alpha(2)-adrenoceptor>dopamine D1 receptor>beta-adrenoceptor, muscarinic and 5-HT(2A/C) receptors. In conscious rats DR4004 (1, 5 or 10 mg/kg i.p.) produced a dose-dependent hyperglycaemia and hypothermia, but the former was reduced by the dopamine D2 receptor antagonist raclopride. Another 5-HT(7) receptor antagonist, (R)-3-(2-(2-(4-methylpiperidin-1-yl)-ethyl)pyrrolidine-1-sulfonyl)phenol (SB-269970) produced hypothermia but no hyperglycaemia. This study confirms that DR4004 has high affinity for the 5-HT(7) receptor but suggests that dopamine D2 receptor activity contributes to some of the in vivo effects.

The synthetic cathinone derivative, mephedrone, is a controlled substance across Europe. Its effects have been compared by users to 3,4-methylenedioxymethamphetamine (MDMA), but little data exist on its pharmacological properties. This study compared the behavioural and neurochemical effects of mephedrone with cathinone and MDMA in rats. Young-adult male Lister hooded rats received i.p. cathinone (1 or 4 mg/kg), mephedrone (1, 4 or 10mg/kg) or MDMA (10mg/kg) on two consecutive days weekly for 3 weeks or as a single acute injection (for neurochemical analysis). Locomotor activity (LMA), novel object discrimination (NOD), conditioned emotional response (CER) and prepulse inhibition of the acoustic startle response (PPI) were measured following intermittent drug administration. Dopamine, 5-hydroxytryptamine (5-HT) and their major metabolites were measured in striatum, frontal cortex and hippocampus by high performance liquid chromatography 7 days after intermittent dosing and 2h after acute injection. Cathinone (1, 4 mg/kg), mephedrone (10mg/kg) and MDMA (10mg/kg) induced hyperactivity following the first and sixth injections and sensitization to cathinone and mephedrone occurred with chronic dosing. All drugs impaired NOD and mephedrone (10mg/kg) reduced freezing in response to contextual re-exposure during the CER retention trial. Acute MDMA reduced hippocampal 5-HT and 5-HIAA but the only significant effect on dopamine, 5-HT and their metabolites following chronic dosing was altered hippocampal 3,4-dihydroxyphenylacetic acid (DOPAC), following mephedrone (4, 10mg/kg) and MDMA. At the doses examined, mephedrone, cathinone, and MDMA induced similar effects on behaviour and failed to induce neurotoxic damage when administered intermittently over 3 weeks.

Methylphenidate (Ritalin) is a selective dopamine reuptake inhibitor and an effective treatment for attention deficit hyperactivity disorder (ADHD) however the anatomical foci and neuronal circuits involved in these therapeutic benefits are unclear. This study determines the temporal pattern of brain regional activity change produced by systemic administration of a therapeutically relevant dose of methylphenidate in anaesthetised Sprague-Dawley rats using BOLD MRI and a 2.35T Bruker magnet. Following 60 min basal recording separate rats received saline (n = 9) or +/- methylphenidate hydrochloride (2 mg/kg, i.p., n = 9) and BOLD changes were recorded for 90 min using statistical parametric maps. Methylphenidate produced significant positive random BOLD effects in the nucleus accumbens, substantia nigra, entorhinal cortex and medial orbital cortex. Negative random BOLD effects were more widespread and intense, occurring in the motor and somatosensory cortices, caudate putamen, lateral globus pallidus and bed nucleus of the stria terminalis, without accompanying changes in blood pressure or respiratory rate. Methylphenidate-induced negative BOLD in the striatum, and other dopamine terminal areas, may reflect post-synaptic changes produced by blockade of the neuronal dopamine reuptake transporter. While increased positive BOLD in the medial orbital cortex may reflect altered dopamine and/or noradrenaline release indirectly altering striatal activity. The overall pattern of BOLD changes is comparable to that seen in previous studies using guanfacine, amphetamine and atomoxetine, and suggests that although these compounds operate through distinct pharmacological mechanisms the BOLD changes may represent a 'fingerprint pattern' predictive of therapeutic benefit in ADHD.

Attention deficit hyperactivity disorder (ADHD) is a common neuropsychiatric disorder characterised by excessive levels of hyperactivity, inattentiveness and impulsivity. Stimulant drugs which increase dopamine neurotransmission are treatments for ADHD. Hypodopaminergic fronto-striatal function with associated overactivity of the dopamine transporter (DAT) represents one possible neurobiological mechanism underlying ADHD. Few, if any, of the existing animal models of ADHD mimic the underlying neurobiology of the disorder. In this study we have further characterised the behavioural profile of a model of a hyperactive inattentive animal through manipulation of the DAT. The behavioural effects of acute treatment and following withdrawal from sub-chronic treatment with GBR 12909 (30 mg/kg i.p.), a potent and highly selective DAT inhibitor, were examined in juvenile rats. GBR 12909 treatment was used to produce a compensatory up regulation following withdrawal. Acute treatment with GBR 12909 (30 mg/kg i.p.) resulted in a marked increase in locomotor and rearing behaviours on the first and fourth days during a 4 consecutive bi-daily drug treatment regime in postnatal weaned rats. Adolescent rats after 10, 20 and 30 days withdrawal from GBR 12909 pre-treatment maintained mild increases in locomotor activity and failed to discriminate a familiar over a novel object in the novel object discrimination task (using both 1 min and 3 h inter-trial intervals) indicating impaired learning and memory. Prepulse inhibition of acoustic startle was unaltered following withdrawal from GBR 12909 treatment. These data reinforce the potential role of the DAT in the underlying neurobiology of ADHD. They also add further evidence to suggest that postnatal changes in the DAT following withdrawal from treatment with the DAT inhibitor, GBR 12909, may prove to be a useful animal model of ADHD with potential for examining the effectiveness of novel ADHD treatments.

The behavioral effects and the biochemical changes produced following either a single or repeated intrathecal injection respectively, of the insect peptide proctolin (Arg-Tyr-Leu-Pro-Thr-OH) have been compared with the effects of a stable analogue of thyrotrophin-releasing hormone (TRH) in rats. Intrathecal proctolin (1-100 micrograms) did not produce any marked behavioural effects on its own, while intrathecal TRH analogue (RX 77368, 0.5 microgram) administration produced wet-dog shakes and forepaw-licking behaviours. Proctolin (10 micrograms) significantly attenuated the wet-dog shake and forepaw-licking behaviours evoked by intrathecal RX 77368 administration when it was given 30 min before, but not when given in combination with RX 77368. Repeated intrathecal proctolin administration (10 micrograms twice daily for 5 days) significantly reduced the 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid and TRH levels in the ventral, but not in the dorsal, horn of the spinal cord nor in the brainstem, and elevated hypothalamic TRH without affecting plasma free thyroxine levels when compared with values in saline-treated controls. Repeated proctolin injection did not alter substance P levels in any brain region examined, nor did it affect the choline acetyltransferase activity or the calcitonin gene-related peptide-like immunoreactive levels in the ventral horn of the spinal cord, both of which are principally located in motoneurones in this cord region.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract Glutamate receptor antagonists have been widely used to study behavioral and neural abnormalities relevant to schizophrenia. We review which human behaviors are affected by these drugs and how relevant they are to the symptoms of schizophrenia and psychosis. We summarize how N-methyl- d -aspartate receptor (NMDA-R) antagonists are considered to induce these behavioral changes and discuss how accurately the acute NMDA-R antagonism model in rodents translates to human conditions. Using a receptor occupancy approach, we test the hypothesis that consideration of pharmacokinetics and pharmacodynamics of NMDA-R antagonists in published studies may help to explain some of the differences between animal and human experiments.