Ondansetron results in improved auditory gating in DBA/2 mice through a cholinergic mechanism

Abstract

The 5-HT₃ receptor antagonist, ondansetron, has been shown to correct the auditory gating deficit in medicated schizophrenia patients. Inhibition of 5-HT₃ receptors releases acetylcholine, the endogenous ligand for nicotinic acetylcholine receptors. The schizophrenia-related auditory gating deficit is modulated, in part, by nicotinic acetylcholine receptors, as is the mouse (DBA/2) model of the deficit. The present study assessed the effects of both acute and chronically administered ondansetron on auditory gating in DBA/2 mice. Auditory gating is defined as a decrease in amplitude of response to the second of a paired identical auditory stimulus presented 0.5 s following an initial auditory stimulus. Acute ondansetron administration at the lowest dose (0.1 mg/kg, IP) tested had no effect, while other doses (0.33 and 1 mg/kg, IP) produced improvements in auditory gating. The improvements were produced through both an increase in response to the first auditory stimulus and a decrease in the response to the second auditory stimulus. Co-administration of an α7 nicotinic acetylcholine receptor antagonist, α-bungarotoxin, or the α4β2 nicotinic acetylcholine receptor antagonist dihydro-β-erythroidine, with the 0.33 mg/kg dose of ondansetron blocked the improvement in auditory gating produced by ondansetron alone. There was no difference in response between the chronically injected mice and naive mice. Both showed improved auditory gating, thus, demonstrating no “carry over” effect of daily injections. These data demonstrate that indirect stimulation of nicotinic acetylcholine receptors by ondansetron can improve auditory gating parameters in DBA/2 mice.

Introduction

The serotonergic and cholinergic neurotransmitter systems have both been implicated in the pathophysiology of schizophrenia (Keshavan et al., 2008). The serotonergic system has been of interest since the 1950s after observations that the psychotomimetic effects of LSD, a serotonin receptor partial agonist, produced clinical effects resembling those of schizophrenia (Fischman, 1983, Woolley and Shaw, 1954). Later studies determined a decrease in 5-HT_2A receptors in the prefrontal cortex of postmortem brain from subjects with schizophrenia (Burnet et al., 1996, Dean and Hayes, 1996, Laruelle et al., 1993) as well as decreased cortical 5-HT_2A receptor mRNA (Burnet et al., 1996, Hernandez and Sokolov, 2000). The majority of serotonin receptors belong to a superfamily of G-protein coupled receptors; however, the 5-HT₃ serotonin receptor is a ligand-gated ion channel.

The 5-HT₃ receptors belong to the same superfamily of ligand-gated ion channels as the nicotinic acetylcholine receptors. There is a 30% sequence homology between the 5-HT₃ receptor and the α7 nicotinic acetylcholine receptor, possessing the greatest sequence similarity within this superfamily of ion channels (Maricq et al., 1991). Like the 5-HT₃ receptor, nicotinic acetylcholine receptors are implicated in the pathophysiology of schizophrenia. Postmortem brain tissue from schizophrenia patients exhibit a decreased density of α7 nicotinic acetylcholine receptors in the CA3 region of the hippocampus and the dentate gyrus as compared to tissue from non-schizophrenia subjects (Breese et al., 1997, Freedman et al., 1995). Furthermore, an endophenotype of the illness, an auditory gating deficit, is linked genetically to chromosome 15q14, which is the locus of the α7 nicotinic acetylcholine receptor gene (Chini et al., 1994, Freedman et al., 1997, Leonard et al., 2002, Orr-Urtreger et al., 1995).

The auditory gating deficit in schizophrenia patients manifests as an inability to inhibit, or filter, the responses to repetitive auditory stimuli. In normal subjects, the ability to filter repetitive auditory information allows for focused attention, while schizophrenia patients report “difficulties in maintaining attention and complain of intrusion of unwanted sensory information” (Adler et al., 1999). Auditory gating is modulated by both the cholinergic and serotonergic systems, among others. The interaction of the serotonergic and cholinergic systems occurs via antagonism of serotonergic receptors. Specifically, antagonism of serotonin 5-HT₃ receptors facilitates the release of acetylcholine, purportedly through disinhibition of GABAergic interneurons (Fink and Gothert, 2007). The release of GABA may correct a deficient inhibitory circuit in the hippocampus thus improving the auditory gating deficit (Adler et al., 1998). The increased release of acetylcholine following blockade of 5-HT₃ receptors should activate nicotinic acetylcholine receptors, thus improving auditory gating. Therefore, the interaction of these neurotransmitter systems can produce various outcomes on auditory gating.

Auditory gating is assessed by recording the electrical responses of the brain to two identical auditory stimuli separated by 500 ms. In control subjects the amplitude of the response to the second stimulus (test) is smaller than the amplitude of the response to the first stimulus (conditioning). However, in schizophrenia patients the response to the test stimulus is of similar magnitude to the response to the conditioning stimulus (Adler et al., 1982).

The equivalent of the human P50 gating measure in rodents is the P20–N40 waveform complex (Bickford-Wimer et al., 1990, Bickford and Wear, 1995, Simpson and Knight, 1993, Stevens et al., 1998). Although it has been proposed that the P20 and N40 waveform responses vary independently according to the pharmacologic or behavioral manipulation (Connolly et al., 2004, Maxwell et al., 2004, Metzger et al., 2007, Umbricht et al., 2004), the entire complex has been shown to have less variability than either the P20 or N40 component alone (Hashimoto et al., 2005). In rodents the P20–N40 complex has been localized to the dorsal hippocampal CA3 region (Bickford-Wimer et al., 1990). The DBA/2 inbred mouse strain is a model for the auditory gating deficit, because (1) it spontaneously fails to inhibit responses to repetitive auditory stimuli (Stevens et al., 1996), (2) the expression level of α7 nicotinic acetylcholine receptors in the hippocampus of DBA/2 mice is decreased as compared to mice with normal auditory gating (Adams et al., 2001) and (3) activation of α7 receptors produce an improvement in the auditory gating deficit of DBA/2 mice (Stevens et al., 1998). Recently, another subtype of nicotinic acetylcholine receptor, α4β2, has also been implicated in the auditory gating deficit in the DBA/2 mouse model (Metzger et al., 2007, Radek et al., 2006, Wildeboer and Stevens, 2008). Therefore, the DBA/2 mouse serves as a relevant model for studying abnormalities in nicotinic receptor function in relation to deficient auditory gating.

Two 5-HT₃ antagonists that produce improvements in auditory gating are tropisetron and clozapine. Tropisetron is an antiemetic and anti-nausea drug used by patients receiving chemotherapy. This compound improves auditory gating in both DBA/2 mice and in subjects with schizophrenia (Hashimoto et al., 2005, Koike et al., 2005). Tropisetron, however, is not only a 5-HT₃ antagonist, but is also a partial agonist at the α7 nicotinic acetylcholine receptor (Macor et al., 2001, Papke et al., 2004, Simpson et al., 2000). Thus, the improvement in auditory gating by tropisetron is proposed to be primarily through the α7 receptor (Hashimoto et al., 2005). Clozapine has also been shown to improve auditory gating in both schizophrenia patients and DBA/2 mice (Nagamoto et al., 1996, Simosky et al., 2003). While clozapine lacks direct agonist activity at the α7 receptor, the study by Simosky and colleagues (2003) demonstrated that the improvement in auditory gating with clozapine was produced by activation of nicotinic receptors, presumably through the increased release of hippocampal acetylcholine (Shirazi-Southall et al., 2002). However, it should be noted that clozapine also antagonizes other neuronal receptors including dopaminergic receptors (Fjalland and Boeck, 1978). Another 5-HT₃ antagonist, ondansetron, improves the auditory gating deficit in medicated schizophrenia patients (Adler et al., 2005). Like clozapine, ondansetron does not have a physiologically relevant binding affinity for α7 or α4β2 nicotinic acetylcholine receptors (Macor et al., 2001) nor does it display direct agonist activity for nicotinic receptors (Papke et al., 2004). Ondansetron has not yet been tested in DBA/2 mice. The purpose of this study was to determine if ondansetron produces an improvement in auditory gating in DBA/2 mice. We hypothesized that ondansetron would improve auditory gating in DBA/2 animals and that the mechanism of improvement would be via nicotinic acetylcholine receptors.