Distribution of 5-HT4 receptors in the postmortem human brain—an autoradiographic study using [125I]SB 207710

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Abstract

The autoradiographic distribution of the 5-HT4 receptor was described using human postmortem brain sections and the selective radioligand [125I]SB 207710 [(1-n-butyl-4-piperidinyl)methyl-8-amino-7-[125I]iodo-1,4-benzodioxane-5-carboxylate]. The specific binding was highest in regions of the basal ganglia (caudate nucleus, putamen, nucleus accumbens, globus pallidus and substantia nigra) and the hippocampal formation (CA1 and subiculum). In the neocortex, the binding showed a distinct lamination pattern with high levels in superficial layers and a band displaying lower levels in deep cortical layers. The results confirm previous studies on the distribution of 5-HT4 receptors in the human brain in vitro and provide high-resolution correlates for in vivo imaging studies using the radioligand recently developed for single photon emission tomography (SPET), [123I]SB 207710.

Introduction

The 5-HT4 receptor has been identified in various tissues as a G-protein coupled receptor positively linked to adenylyl cyclase activity (Hoyer et al., 1994). The presence of 5-HT4 receptors has been described in the brain of several species (Domenech et al., 1994, Jakeman et al., 1994), including human (Bonaventure et al., 2000, Domenech et al., 1994, Mengod et al., 1996, Reynolds et al., 1995, Waeber et al., 1993). High densities of 5-HT4 receptors are found in regions of the basal ganglia with lower levels in the hippocampal formation and the neocortex (Bonaventure et al., 2000, Domenech et al., 1994, Grossman et al., 1993, Jakeman et al., 1994, Reynolds et al., 1995, Waeber et al., 1993, Waeber et al., 1994). In the human brain, 5-HT4 receptor mRNA levels are high in the hippocampal formation and in the striatum but are, in contrast to the receptor proteins, absent in the globus pallidus and the substantia nigra, indicating that 5-HT4 receptors are localised in terminals of striatal projections to these regions (Bonaventure et al., 2000).

While 5-HT4 receptors in the alimentary tract are suggested to be important targets for the treatment of gastrointestinal motility disorders (De Ponti and Tonini, 2001), the function of central 5-HT4 receptors is less well established. However, the distribution pattern in combination with neurochemical and behavioural observations indicates that brain 5-HT4 receptors may be involved in cognition and anxiety (Eglen et al., 1995).

There are a few reports comparing brain 5-HT4 receptor densities postmortem in subjects with neuropsychiatric disorders with those of normal control subjects. Reynolds et al. (1995) observed lower levels of 5-HT4 receptors in the putamen of brains of patients affected by Huntington’s disease and also reduced 5-HT4 receptor densities in the hippocampus and frontal cortex in Alzheimer’s disease patients compared to controls. On the other hand, no differences were found in the brains from parkinsonian patients (Reynolds et al., 1995). Furthermore, no differences in 5-HT4 receptor densities could be detected in the dorsolateral prefrontal cortex of brains from schizophrenic patients compared to controls (Dean et al., 1999).

In the present study, we have used the 5-HT4 receptor antagonist radioligand [125I]SB 207710 [(1-n-butyl-4-piperidinyl) methyl-8-amino-7-[125I]iodo-1,4-benzodioxane-5-carboxylate] (Brown et al., 1993) to examine the distribution of 5-HT4 receptors in the postmortem human brain by autoradiography. Binding studies indicate that this compound shows high affinity for 5-HT4 receptors (KD values of 86 and 37 pM in piglet hippocampus and caudate membranes, respectively) and about 1000-fold selectivity compared to other 5-HT receptors (Brown et al., 1993). [125I]SB 207710 has also been used in autoradiographic studies to visualise brain 5-HT4 receptors in rat (Patel et al., 1995, Vilaro et al., 1996) and several other species, including human (Mengod et al., 1996).

Iodine-123 labelled SB 207710 has recently been shown to be an effective radioligand for single photon emission tomography (SPET) studies in vivo (Pike et al., 1999). Thus, in a SPET investigation of the cynomolgus monkey brain, [123I]SB 207710 showed suitable in vivo characteristics and was accumulated in 5-HT4 receptor-rich regions including the striatum and the neocortex (Pike et al., 1999). This preliminary finding reinforces further characterisation of SB 207710 as radioligand for the study of 5-HT4 receptors in the human brain in vivo. In the present study, we used human postmortem brain whole hemisphere autoradiography (Hall et al., 1998, Hall et al., 2001) to provide detailed high resolution anatomical correlates for the lower resolution in vivo studies.

Section snippets

Compounds

[125I]SB 207710 was prepared from the precursor SB 207715 [(1-n-butyl-4-piperidinyl)methyl-8-amino-7-tributylstannyl-1,4-benzodioxane-5-carboxylate]. SB 207715 was incubated in [125I]sodium iodide solution (no-carrier-added) in the presence of chloramine-T. The radioligand (specific radioactivity, 2200 Ci/mmol) was purified on a reverse phase column (μ-Bondapak C18), evaporated and dissolved in 75% ethanol. The radiochemical yield was 60% and the radiochemical purity was over 99%. Formulated [

General

The general brain distribution of [125I]SB 207710 is shown in the autoradiograms obtained using coronal (Fig. 1) and horizontal (Fig. 2) sections. High levels of [125I]SB 207710 binding sites were observed in regions of the basal ganglia and the hippocampus with lower levels in the neocortex. Generally, the non-specific binding of [125I]SB 207710, as determined in the presence of 10 μM serotonin, was relatively low. Accumulation of radioactivity in the white matter was relatively high and was

Discussion

This study shows that [125I]SB 207710 is suitable for the autoradiographic mapping of 5-HT4 receptors in human brain whole hemisphere sections. Studies using [125I]SB 207710 are therefore well suited to provide detailed anatomical correlates for low-resolution SPET images obtained with [123I]SB 207710 in vivo.

The localisation of 5-HT4 receptors in the human brain has been described using a similar autoradiographic methodology and the radioligands [3H]prucalopride and [3H]R116712 (Bonaventure et

Acknowledgments

The excellent technical assistance of Siv Eriksson, David Card, Dr Rachel S. Mulligan and Kerstin Larsson is greatly acknowledged. We are grateful for the discussions on neuroanatomy with Professor Yasmin L. Hurd. This work was supported by the Human Brain Informatics (HUBIN) at Karolinska Institutet and by grants from the Swedish Medical Research Council (MFR) 11640 and the National Institute of Mental Health (NIMH) MH 44814. Parts of the results have been presented in abstract form at the

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