Long-Term Administration of Citalopram Reduces Basal and Stress-Induced Extracellular Noradrenaline Levels in Rat Brain

Psychopharmacology (2007) 194:73–81 DOI 10.1007/s00213-007-0826-8 ORIGINAL INVESTIGATION Long-term administration of citalopram reduces basal and stress-induced extracellular noradrenaline levels in rat brain Yukie Kawahara & Hiroshi Kawahara & Fumi Kaneko & Masatoshi Tanaka Received: 24 December 2006 /Accepted: 7 May 2007 /Published online: 30 May 2007 # Springer-Verlag 2007 Abstract suppressed only in the LC. The effect of local application of Rationale Panic disorders are commonly treated with clonidine was enhanced only in the BLA. selective serotonin reuptake inhibitors (SSRIs). However, Conclusion The present results indicate that chronic ad- the effect of SSRIs on noradrenaline systems in the brain ministration of citalopram strongly decreases the extracel- has not been fully elucidated at the present time. lular levels of noradrenaline in the brain. The anti-panic Objectives The effects of long-term administration of effect of citalopram might be due to sensitization of the α2- citalopram, an SSRI, on basal as well as stress-induced adrenoceptors leading to suppression of the stress response extracellular noradrenaline levels in the basolateral nucleus through noradrenergic activity. This mechanism is specific of the amygdala (BLA) and the locus coeruleus (LC) were for the BLA. determined. In addition, the responsiveness of noradrenaline transporters and α2-adrenoceptors were determined Keywords Citalopram . Microdialysis . Amygdala . after long-term administration of citalopram. Basolateral . Rat . Noradrenaline . Chronic treatment . Materials and methods Brain microdialysis was used to Locus coeruleus assess the extracellular levels of noradrenaline in conscious rats. Desipramine and clonidine were used to functionally evaluate the noradrenaline transporter and α2-adrenoreceptor, Introduction respectively. Results In rats treated daily for 14 days with citalopram Dysfunction of the central noradrenergic system has been −1 −1 (10 mg kg day s.c.), dialysate noradrenaline levels demonstrated in anxiety disorders, such as panic disorders, showed remarkable decreases in both the BLA and the LC in various studies (Brunello et al. 2003; Ressler and to about 25 and 45% of controls, respectively. The stress- Nemeroff 2001; Sullivan et al. 1999). The involvement of induced increase of noradrenaline was almost completely noradrenergic systems in stress responses, in particular in abolished in the BLA, but was relatively stable in the LC. the amygdala, locus coeruleus and hypothalamus, has been The effect of local application of desipramine tended to be reported by ourselves and others (Debiec and LeDoux 2006; Neophytou et al. 2001; Tanaka et al. 2000). As general activation of these brain regions seem to be involved in the pathology of panic disorders, it is of interest * : : Y. Kawahara ( ) F. Kaneko M. Tanaka to investigate whether selective serotonin reuptake inhibitor Department of Pharmacology, Kurume University School of Medicine, (SSRI) might affect the activity of noradrenergic neurons in Kurume 830 0011, Japan these regions (Gorman et al. 2000). e-mail: [email protected] Despite the fact that the pathology of anxiety disorders has been related to the noradrenergic system, SSRIs such as H. Kawahara Department of Dental Anesthesiology, Kyushu Dental College, citalopram, fluoxetine, and proxetine, are currently the first Kitakyushu 803 8580, Japan line treatment of anxiety disorders, i.e., panic and obses- 74 Psychopharmacology (2007) 194:73–81 sive-compulsive disorders (Tukel et al. 2006; Seedat et al. serotonin transporters, with little affinity for noradrenaline 2003; Pollock 2001; Wade 1999). transporters (Owens et al. 1997). The noradrenaline It has been well established that SSRIs selectively block transporter and α2-adrenoceptors have been reported to be serotonin reuptake sites (Owens et al. 1997), and it is desensitized after chronic treatment with noradrenaline assumed that their clinical effects are attributed to increased reuptake inhibitors (Benmansour et al. 2004; Invernizzi levels of serotonin in the brain. However, accumulating and Garattini 2004). Therefore, we investigated whether evidence indicates that the noradrenergic system is also chronic citalopram affected the noradrenaline transporters implicated in the action of SSRIs. For instance, extracellu- and α2-adrenoceptors. For this purpose, the effects of local lar noradrenaline levels in the cortex are increased by infusion of desipramine and clonidine into the BLA or the SSRIs when administered both acutely and repeatedly LC by retrograde microdialysis in the chronically citalopram- (David et al. 2003; Thomas et al. 1998). Electrophysiolo- treated group were compared with saline-injected controls. gical studies have demonstrated that repeated administration of citalopram, as well as paroxetine, reduced the spontaneous firing rate of noradrenergic neurons (Szabo et al. 1999, 2000). In humans, significant normalization of Materials and methods noradrenergic function has been demonstrated during effective treatment with SSRIs in patients with panic Animals disorder in which a greater reduction of the major metabolite of noradrenaline, 3-methoxy-4-hydroxyphenyle- Male albino Wistar rats (280–320 g) were used for these thyleneglycol (MHPG) was observed compared to healthy experiments. The rats were maintained at 23±2°C under a volunteers (Coplan et al. 1997). Although the interaction of 12-h light–dark cycle (lights on at 08:00 A.M.) with free noradrenaline and serotonin has been demonstrated previ- access to food and water. After probe implantation, the rats ously (Svensson 2000; Shiekhattar and Aston-Jones 1993), were housed individually in plastic cages (30×30×40 cm). the mechanism by which SSRIs interact with the noradrenergic system is largely unknown. Based on these reports, Materials we presumed that the noradrenergic system could be functionally altered when serotonergic neuronal activity is 1-(3-Dimethylaminopropyl)-1-(4-fluorophenyl)-5-phtalan- enhanced by long-term administration of SSRIs. carbonitril (citalopram hydrobromide, generously provided The basolateral amygdaloid nucleus (BLA) serves as an by H. Lundbeck A/S, Copenhagen, Denmark) was dissolved integrator and relay center for the sensory and memory in saline (0.2 ml) or Ringer’s solution for systemic injection information necessary for anxiety and panic responses and local application, respectively. (Shekhar et al. 2003). Neuroimaging studies indicate that the amygdala is involved in the provocation of fear (Morris Surgery and brain dialysis et al. 1999). The BLA receives input from both noradrenergic and serotonergic neurons mainly from the brain stem, Microdialysis was performed with an I-shaped cannula. The e.g., A6 and B9 cell groups, respectively, and these neurons probe was implanted in the right BLA (exposed length are highly responsive to various types of aversive stimuli 1.0 mm) or the right LC (exposed length 1.5 mm) under (Uehara et al. 2005; Ferry and McGaugh 2000). Therefore, pentobarbital anesthesia (50 mg/kg i.p.) and local applica- it is likely that SSRIs modulate these neuronal inputs to the tion of 10% lidocaine. The coordinates of the implantation BLA, and this effect could be involved in the clinical were: A/P −2.8 mm, L/M 5.0 mm, V/D 8.8 mm from the efficacy of SSRIs in anxiety disorders. In addition, we bregma and dura for the BLA. For the LC, A/P −3.3 mm, selected the locus coeruleus (LC) to determine noradren- L/M 1.3 mm, V/D 8.3 mm at the angle of 15° in the sagittal aline levels, which may reflect a general effect of SSRIs on plane from lambda and the surface of the scull, respectively. the noradrenergic system, as most of the noradrenaline in Microdialysis experiments were conducted 24 h after the central nervous system is derived from the LC. implantation of the probes as described in previous reports The present microdialysis study was designed to studying noradrenaline (McKittrick and Abercrombie 2007; characterize the effect of long-term administration of Valentini et al. 2005) An online approach was used in citalopram on extracellular levels of noradrenaline in the which the probes were perfused with Ringer’s solution at a BLA and the LC. The effect of handling stimuli, which is flow rate of 2.0 ml/min by an infusion pump (EICOM, considered to elicit a mild emotional stressful state, on the Kyoto, Japan). Dialysate fractions were collected every release of noradrenaline was determined after long-term 20 min. administration of citalopram. Citalopram was selected, as it Noradrenaline was quantified by high-performance has the greatest selectivity among the known SSRIs for liquid chromatography using a reverse-phase column Psychopharmacology (2007) 194:73–81 75 Table 1 Basal noradrenaline levels in dialysates: control vs long-term Results citalopram Control Long-term citalopram Effect of long-term administration of citalopram on basal levels of noradrenaline BLA 5.36±1.33 (n=16) 1.33±0.19 (n=21)** n n LC 4.62±0.81 ( =19) 2.04±0.43 ( =14)* The mean basal values for noradrenaline in the BLA and Data are expressed as mean±SEM (fmol/20 min). the LC corresponding to each experimental group are *Indicates p<0.05 provided in Table 1. **Indicates p<0.01 vs control The noradrenaline levels were significantly reduced, compared to baseline, after long-term administration of (150×4.6 mm; Supelco LC18, Belleofonte, PA, USA) with citalopram in both the BLA (to about 25% of controls) and electrochemical detection. An EICOM EP-300 pump (Kyoto, the LC (to about 45% of controls) as summarized in Fig. 1. Japan) was used in conjunction with an electrochemical detector (ESA; potential first cell, +180 mV; potential second Effect of long-term administration of citalopram cell, −180 mV). The mobile phase consisted of a mixture of on stress-induced noradrenaline in dialysates 4.1gsodiumacetateadjustedtopH5.5,50mgNa2EDTA and Handling for 20 min caused a rapid and significant increase of 140 mg octanesulfonic acid in 900 ml H2O, and 100 ml/ l methanol. The flow rate was 1.0 ml/min. The detection limit noradrenaline to about 200% of basal levels in both the BLA of the assay was about 0.3 fmol per sample (on-column).

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