Tsukasa Koyama

Effects of conditioned fear stress on serotonin neurotransmission and freezing behavior in rats

In an attempt to clarify the role of the brain serotonergic system in the psychopathology of anxiety, we examined the effect of a psychological stress, conditioned fear stress, on extracellular serotonin (5-hydroxytryptamine, 5-HT) concentrations in the rat medial prefrontal cortex using the method of in vivo microdialysis, while simultaneously observing conditioned fear stress-induced freezing behavior, an index of anxiety. Conditioned fear stress increased extracellular 5-HT levels in the medial prefrontal cortex, and this 5-HT level increase was followed by a resolution of the freezing behavior. A dose of 10 mg/kg of a selective 5-HT reuptake inhibitor, citalopram, administered 60 min before exposure to conditioned fear stress increased extracellular 5-HT concentrations immediately and potently, reducing freezing behavior. These findings strongly suggest that facilitation of brain 5-HT neurotransmission decreases anxiety, which is in agreement with the clinical reports that selective 5-HT reuptake inhibitors are effective in the treatment of anxiety disorders.

Effect of the Dopamine D1/5 Antagonist SCH 23390 on the Acquisition of Conditioned Fear

The authors previously reported that typical and atypical antipsychotic drugs inhibited the acquisition but not expression of conditioned fear. The present study examined the effects of the selective dopamine D(1/5) agonist (SKF 38393) and antagonist (SCH 23390) on the acquisition and expression of conditioned fear. Drugs were administered subcutaneously to male Sprague-Dawley rats 30 min before foot shock (2.5 mA for 5 min). Twenty-four hours after foot shock, rats were again placed and observed in the shock chamber without shocks (conditioned fear). Freezing behavior induced by conditioned fear, an index of anxiety or fear, was recorded using a time-sampling procedure. SCH 23390 (0.1-1 mg/kg) inhibited the acquisition of conditioned freezing. The administration of SCH 23390 at a dose of 0.1 mg/kg 30 min after foot shock did not affect conditioned freezing. Taken together, it is concluded that D(1/5) antagonism inhibits the acquisition of conditioned fear. SKF 38393 (3-20 mg/kg) failed to change the acquisition of conditioned fear. SCH 23390 or SKF 38393 administered prior to testing did not reduce the expression of conditioned fear. These results suggest that D(1/5) receptors may play a role in the development of fear or anxiety.

Effect of chronic administration of flesinoxan and fluvoxamine on freezing behavior induced by conditioned fear

The present study investigated the acute effects of flesinoxan (a selective 5-HT(1A) receptor agonist), fluvoxamine (a selective serotonin reuptake inhibitor) and their co-administration on the expression of conditioned freezing, and index of anxiety in rats. This study also examined the acute effects of fluvoxamine and flesinoxan following chronic flesinoxan or chronic fluvoxamine on the expression of conditioned freezing. Acute administration of flesinoxan (s.c.; 0.1-3 mg/kg) reduced freezing dose dependently, and fluvoxamine (i.p.) at a high dose (60 mg/kg) reduced freezing significantly. Acute co-administration of fluvoxamine (30 mg/kg) and flesinoxan (0.3 mg/kg) showed an additive inhibitory effect on freezing. Chronic flesinoxan treatment (0.3 mg/kg, for 13 days) did not affect the inhibitory effect of acute flesinoxan treatment, but enhanced that of acute fluvoxamine (30 mg/kg) on conditioned freezing. Chronic fluvoxamine treatment (30 mg/kg, for 13 days) enhanced the inhibitory effect of acute fluvoxamine (30 mg/kg) and the inhibitory effect of acute flesinoxan (0.3 mg/kg) on conditioned freezing. These results suggest that co-administration of a selective serotonin reuptake inhibitor and a 5-HT(1A) receptor agonist is useful for the treatment of anxiety disorders.

Monoamine oxidase inhibitors reduce conditioned fear stress-induced freezing behavior in rats

The present study examined the acute anxiolytic effects of monoamine oxidase inhibitors on freezing behavior, a putative index of anxiety induced by conditioned fear stress. The selective serotonin 1A receptor agonist tandospirone (0.1-10 mg/kg) inhibited freezing dose dependently. The irreversible, non-selective monoamine oxidase inhibitors tranylcypromine (3 and 15 mg/kg) and phenelzine (30 and 80 mg/kg) reduced freezing significantly. Clorgyline (10 mg/kg, irreversible selective monoamine oxidase A inhibitor), N-(2-aminoethyl)-5-(m-fluorophenyl)-4-thiazole carboxamide (Ro 41-1049) (30 mg/kg, reversible selective monoamine oxidase A inhibitor), selegiline (3 mg/kg, irreversible selective monoamine oxidase B inhibitor) and lazabemide (10 mg/kg, reversible selective monoamine oxidase B inhibitor) had no effect on freezing behavior. However, combined administration of clorgyline (10 mg/kg) and selegiline (3 mg/kg) reduced freezing significantly, as well as combined administration of clorgyline (10 mg/kg) and lazabemide (10 mg/kg), Ro 41-1049 (30 mg/kg) and selegiline (3 mg/kg), or Ro 41-1049 (30 mg/kg) and lazabemide (10 mg/kg). These effects of monoamine oxidase inhibitors on freezing were not due to non-specific motor effects. These results suggest that acute inhibition of both monoamine oxidase A and B reduced anxiety or fear, while inhibition of monoamine oxidase A or B alone failed to reduce anxiety or fear.

Effect of subchronic lithium treatment on citalopram‐induced increases in extracellular concentrations of serotonin in the medial prefrontal cortex

We investigated the effect of citalopram [a selective serotonin (5‐HT) reuptake inhibitor; SSRI] and MKC‐242 (a selective 5‐HT1A agonist), following treatment with subchronic lithium (p.o., 1 week) on extracellular 5‐HT concentrations in the medial prefrontal cortex (mPFC). Acute treatment with citalopram (3 and 30 mg/kg) led to significant increases in extracellular 5‐HT concentrations. The subchronic lithium group showed significantly higher basal levels of extracellular 5‐HT than normal diet controls. Acute citalopram (3 and 30 mg/kg) treatment together with subchronic lithium treatment showed significant increases in the extracellular 5‐HT concentrations, compared with citalopram treatment alone. Acute MKC‐242 (1 mg/kg) treatment showed significant decreases in extracellular 5‐HT concentrations, in both the normal diet and lithium diet groups to the same extent. The addition of lithium did not change the effect of the 5‐HT1A agonist on extracellular 5‐HT concentrations. This study suggests that lithium augmentation of the antidepressant effect of SSRI is mediated by the additional increases in extracellular 5‐HT concentrations following the co‐administrations of lithium and SSRI.

Effects of the Co-administration of 5-HT1A Receptor Antagonists with an SSRI in Conditioned Fear Stress-Induced Freezing Behavior

The effects of the co-administration of the serotonin (5-HT) 1A receptor antagonists NAN-190 or (+)-WAY100135 with a selective 5-HT reuptake inhibitor (SSRI) citalopram on conditioned fear stress (CFS)-induced freezing behavior, which is the animal model of anxiety, were examined. The inhibitory effects of co-administration of NAN-190 (0.1-10 mg/kg) with citalopram on CFS-induced freezing were potent; in particular, at 0.1 and 0.25 mg/kg, NAN-190 significantly enhanced the effect of citalopram alone. At 0.1 mg/kg, (+)-WAY100135 also markedly enhanced the inhibitory effect of citalopram on freezing behavior. These findings suggest that 5-HT1A receptor antagonist, particularly at low doses, enhances the antifreezing effect of citalopram by blocking the autoreceptor-mediated negative feedback mechanisms of the 5-HT neuron. These experimental results concur with clinical findings that 5-HT1A receptor antagonist pindolol potentiates the effect of 5-HT reuptake inhibitors.

Effect of subchronic lithium carbonate treatment on anxiolytic-like effect of citalopram and MKC-242 in conditioned fear stress in the rat

We investigated the effect of citalopram [selective serotonin (5-HT) reuptake inhibitor] and MKC-242 (5-[3-¿(2S)-(1, 4-Benzodioxan-2-ylmethyl) amino¿propoxy]-1, 3-benzo-dioxol hydrochloride; a selective 5-HT(1A) receptor agonist) on the expression of conditioned freezing, an index of anxiety, following treatment with subchronic lithium carbonate (LiCO(3)). Male Sprague-Dawley rats were used in these experiments. Acute administration of citalopram (subcutaneously, s.c.) reduced freezing significantly at a high dose (30 mg/kg), while showing no effect at lower doses (3 and 10 mg/kg). Acute administration of MKC-242 (s.c.; 0.1-10 mg/kg) dose dependently reduced freezing. Subchronic LiCO(3) treatment (1 week; 0.05% or 0.2% LiCO(3) in diet; p.o.) showed no effect on freezing behavior. Acute treatment with both citalopram (3 and 30 mg/kg) and MKC-242 (1 mg/kg) after subchronic treatment with the higher, but not the lower concentration of LiCO(3) (1 week), reduced freezing markedly and significantly, as compared with either drug alone. These results suggest that subchronic LiCO(3) treatment enhanced the anxiolytic-like effects of these serotonergic drugs by facilitating central 5-HT neurotransmission at clinically therapeutic plasma lithium levels.

Effects of acute citalopram on the expression of conditioned freezing in naive versus chronic citalopram-treated rats.

An acute challenge with selective serotonin (5-HT) reuptake inhibitors (SSRIs) reduces the conditioned freezing in rats, a model of anxiety. The increase in the 5-HT levels in the nerve terminal induced by SSRIs is closely related to its pharmacological effects. Clinically, SSRIs exert an anxiolytic effect after chronic treatment. The effects of repeated treatment with citalopram on conditioned freezing in rats were examined in the present study. Acute citalopram (10 mg/kg) reduced freezing at a short post-training interval (1 day) significantly. While the effect of citalopram (10 mg/kg) on freezing was diminished by prolonging the interval between conditioning and the exposure to conditioned fear stress, repeated citalopram (10 mg/kg) injection twice daily for 7 days restored the inhibitory effect of acute challenge of citalopram (10 mg/kg) on freezing. By prolonging the period between conditioning and exposure to conditioned fear stress, this model may have a more precise predictive validity of anxiety disorder as an animal model.

Effect of co-administration of the selective 5-HT1A receptor antagonist WAY 100,635 and selective 5-HT1B/1D receptor antagonist GR 127,935 on anxiolytic effect of citalopram in conditioned fear stress in the rat.

This study investigated the effect of co-administration of the selective 5-HT1A receptor antagonist WAY 100,635 and selective 5-HT(1B/1D) receptor antagonist GR 127,935 with a subactive dose of citalopram [selective serotonin (5-HT) reuptake inhibitor (SSRI)] on the expression of conditioned freezing, an index of fear. In the present study, acute administration of citalopram (s.c.) reduced freezing significantly at high doses (10, 30 and 100 mg/kg), while showing no significant effect at low doses (1 and 3 mg/kg). Co-administration of WAY 100,635 (0.15 mg/kg) with citalopram (3 mg/kg) reduced freezing markedly and significantly, as compared with either drug alone. However, the addition of GR 127,935 (4 mg/kg) did not potentiate the effects of citalopram (3 mg/kg) on freezing and did not enhance the effect of WAY 100,635 (0.15 mg/kg) with citalopram (3 mg/kg). Co-administration of WAY 100,635 (0.15 mg/kg) or GR 127,935 (4 mg/kg) gave no effect on high-dose citalopram (30 mg/kg)-induced inhibition of freezing behavior. These results suggest that co-administration of WAY 100,635 (0.15 mg/kg) strengthens the anxiolytic effect of citalopram (3 mg/kg) by facilitating central 5-HT neurotransmission. Since GR 127,935 (4 mg/kg) failed to accelerate the inhibition of freezing induced by citalopram (3 mg/kg) with WAY 100,635 (0.15 mg/kg) or citalopram (3 mg/kg) alone, it is suggested that blocking 5-HT1A receptors is more effective in facilitating the anxiolytic effect of citalopram than blocking 5-HT1B/1D receptors.

Synergistic effects of tandospirone and selective serotonin reuptake inhibitors on the contextual conditioned fear stress response in rats

The purpose of this study was to investigate the effects of co-administration of tandospirone (a 5-HT(1A) receptor agonist) and individual selective serotonin reuptake inhibitors (SSRIs) on the contextual conditioned fear stress, using an anxiety model in rats. One day after fear conditioning, tandospirone (0.3-3 mg/kg, s.c.), paroxetine (5-20 mg/kg, i.p.), fluvoxamine (30-60 mg/kg, i.p.) and citalopram (3-30 mg/kg, i.p.) significantly inhibited the conditioned freezing in a dose-dependent manner, whereas, 14 days after fear conditioning, the anxiolytic effects of these drugs were weakened. Fourteen days after fear conditioning, co-administration of tandospirone (0.3 mg/kg, s.c.) with each SSRI [paroxetine (5 mg/kg, i.p.), fluvoxamine (30 mg/kg, i.p.) and citalopram (10 mg/kg, i.p.)], given at subeffective doses, markedly inhibited the conditioned freezing without affecting the locomotor activities and CYP3A4-related pharmacokinetic drug-drug interaction. These results elucidate the pharmacodynamic synergistic effects of tandospirone and SSRIs. Therefore, this augmentation therapy (SSRI+5-HT(1A) receptor agonist) may prove useful for some anxiety disorders.