Ilga Misane

Decreased 5-HT transporter mRNA in neurons of the dorsal raphe nucleus and behavioral depression in the obese leptin-deficient ob/ob mouse.

The neurotransmitter serotonin (5-hydroxytryptamine; 5-HT) is an important regulator of feeding behavior. A hypothalamic site of action for 5-HT in body weight control is supported by the presence of 5-HT receptors in hypothalamic regions which are intimately associated with regulation of food intake. In the present study we have investigated whether there may be an interaction between the hormone leptin, an adipose tissue-derived cytokine signaling factor that inhibits food intake and lowers body weight, and the brain serotonergic system. Immunohistochemical analysis of colchicine-treated rats showed colocalization of 5-HT transporter- and leptin receptor-immunoreactivity in cell bodies of the dorsal raphe nucleus, suggesting that dorsal raphe neurons are targets for circulating leptin. Levels of 5-HT transporter mRNA expression were compared in neurons of the dorsal raphe nucleus of obese leptin-deficient ob/ob mice and their lean littermates using in situ hybridization. 5-HT transporter mRNA levels were significantly down-regulated in neurons of the dorsal raphe nucleus of obese ob/ob mice as compared to lean control mice. Behavioral analysis showed that obese ob/ob mice had significantly lower locomotor activity and exhibited increased immobility in Porsolts test, a model for depression. Taken together, these results suggest that serotonergic cell bodies in the rodent dorsal raphe nucleus possess leptin receptors and that the serotonergic system, as reflected by expression levels of 5-HT transporter mRNA, is down-regulated in the obese behaviorally depressed ob/ob mouse.

A Behavioral Analysis of the Spatial Learning Deficit Induced by the NMDA Receptor Antagonist MK-801 (Dizocilpine) in the Rat

This study analyzes whether the disruptive effects of the noncompetitive NMDA receptor antagonist MK-801 (0.01– 0.1 mg/kg SC) on spatial learning can be dissociated from sensorimotor disturbances in the rat. Two different modifications of the Morris swim maze task with a hidden underwater platform were used: with or without local cue. Retention was tested either 24 h or 7 days after training as a probe trial (without platform). The present data indicate that MK-801 produces an impairment of spatial learning that cannot be dissociated from motor or sensory mechanisms. These findings support the view that NMDA receptors probably contribute to, but are not essential for, spatial learning in the water maze.

Galanin Modulates 5‐Hydroxytryptamine Functions: Focus on Galanin and Galanin Fragment/5‐Hydroxytryptamine1A Receptor Interactions in the Braina

Abstract: The reciprocal interactions between galanin and 5‐HT1A receptors in the rat brain are presented. Galanin and its NH2‐terminal fragments antagonize 5‐HT1A receptor‐mediated transmission at the postjunctional level, whereas galanin receptor activation mimics the inhibitory action of 5‐HT1A receptor activation at the soma‐dendritic level, leading to reductions of 5‐HT metabolism and release. These interactions have been shown in both receptor binding studies and functional studies. In view of the present findings, galanin antagonists may represent a new type of antidepressant drug, based on the 5‐HT hypothesis of depression, by enhancing 5‐HT release and postjunctional 5‐HT1A‐mediated transmission. Moreover, following intracerebroventricular injection galanin was found to be internalized in a population of hippocampal nerve cells mainly representing GABA, somatostatin, and/or NPY‐immunoreactive nerve cells. The relevance of these findings is discussed in relation to the concept of volume transmission.

Modulation of acetylcholine and serotonin transmission by galanin. Relationship to spatial and aversive learning.

Abstract: This paper presents evidence that galanin is a potent in vivo modulator of basal acetylcholine release in the rat brain with qualitatively and quantitatively differential effects in the dorsal and ventral hippocampus. Galanin perfused through the microdialysis probe decreased basal acetylcholine release in the ventral hippocampus, while it enhanced acetylcholine release in the dorsal hippocampus. Galanin (3 nmol/rat) infused into the ventral hippocampus impaired spatial learning acquisition, while it tended to facilitate acquisition when injected into the dorsal hippocampus. These effects appear to be related to activation of GAL‐R1 (ventral hippocampus) and GAL‐R2 (dorsal hippocampus) receptors, respectively. However, the effects of galanin on acetylcholine release and on spatial learning appear not to be directly related to cholinergic mechanisms, but they may also involve interactions with noradrenaline and/or glutamate transmission. Galanin administered into the lateral ventricle failed to affect acetylcholine release, while this route of administration produced a long‐lasting reduction in 5‐HT release in the ventral hippocampus, indicating that galanin is a potent inhibitor of mesencephalic 5‐HT neurotransmission in vivo. Subsequent studies supported this hypothesis, showing that the effects on 5‐HT release in vivo are most likely mediated by a galanin receptor in the dorsal raphe. The implications of these findings are discussed in relation to the role of acetylcholine in cognitive functions in the forebrain and the role of the raphe 5‐HT neurons in affective disorders.

Multiple 5-HT Receptors in Passive Avoidance: Comparative Studies of p-Chloroamphetamine and 8-OH-DPAT

The aim of this study was to examine the involvement of multiple 5-HT receptors in passive avoidance (PA) with a focus on 5-HT1A, 5-HT2A, and 5-HT2C receptors. Because increases in 5-HT transmission result in concomitant multiple 5-HT receptor activation, the effects of the 5-HT releasing compound p-chloroamphetamine (PCA) were compared with those of the selective 5-HT1A receptor agonist 8-OH-DPAT in the rat. In addition, some results with the nonselective 5-HT2C/2B/1B receptor agonist mCPP are presented. When injected before PA training, 8-OH-DPAT, mCPP, and PCA produced a dose-related impairment of the 24-hour retention. The crucial involvement of the postsynaptic 5-HT1A receptors in the action of 8-OH-DPAT was confirmed. Thus, the 5-HT1A receptor antagonists WAY 100635 and (−)-pindolol blocked the PA deficit by 8-OH-DPAT. The impairment of PA caused by PCA was attenuated by WAY 100635 and (−)-pindolol, suggesting an involvement of the 5-HT1A receptor. In contrast, the 5-HT2A and 5-HT2C receptors were of negligible importance in the 24-hour retention deficit induced by PCA. However, the ability of the 5-HT2C receptor antagonist Ro 60-0491 to block the inhibitory effects of mCPP indicated an important regulatory role of the 5-HT2C receptor in PA. The nonselective 5-HT receptor antagonist methiothepin attenuated the PA deficit by PCA but lacked activity versus 8-OH-DPAT. These data provide evidence for the hypothesis that, in addition to the 5-HT1A receptor, other 5-HT receptor subtypes are involved in the inhibitory actions of PCA. Importantly, changes in dopamine transmission seemed not to contribute to the PA impairment by PCA. The behavioral alterations caused by the drug treatments at the time of PA training could not be related to the subsequent retention performance. In conclusion, multiple 5-HT receptors are involved in PA with roles that probably differ at various stages of information processing. These findings also suggest that there probably exists a functional distinction between 5-HT receptor subtypes in different types of aversive learning.

Analysis of the 5-HT1A receptor involvement in passive avoidance in the rat.

1 The effects of the 5‐HT2A/2C agonist DOB, the selective 5‐HT1A agonist NDO 008 (3‐dipropylamino‐5‐hydroxychroman), and the two enantiomers of the selective 5‐HT1A agonist 8‐OH‐DPAT (R(+)‐8‐OH‐DPAT and S(−)‐8‐OH‐DPAT) were studied in a step‐through passive avoidance (PA) test in the male rat. 2 The 5‐HT1A agonists injected prior to training (conditioning) produced a dose‐dependent impairment of PA retention when examined 24 h later. R(+)‐8‐OH‐DPAT was four times more effective than S(−)‐8‐OH‐DPAT to cause an impairment of PA retention. Both NDO 008 and the two enantiomers of 8‐OH‐DPAT induced the serotonin syndrome at the dose range that produced inhibition of the PA response, thus, indicating activation of postsynaptic 5‐HT1A receptors. 3 Neither NDO 008 nor R(+)‐8‐OH‐DPAT induced head‐twitches, a behavioural response attributed to stimulation of postsynaptic 5‐HT2A receptors. In contrast, DOB induced head‐twitches at the 0.01 mg kg−1 dose while a 200 times higher dose was required to produce a significant impairment of PA retention. 4 The impairment of PA retention induced by both NDO 008 and R(+)‐8‐OH‐DPAT was fully blocked by the active S(+)‐ enantiomer of the selective 5‐HT1A antagonist WAY 100135 and the mixed 5‐HT1A/β‐adrenoceptor antagonist L(−)‐alprenolol. In contrast, the mixed 5‐HT2A/2C antagonists ketanserin and pirenperone were found to be ineffective. Moreover, the β2‐adrenoceptor antagonist ICI 118551, the β1‐antagonist metoprolol as well as the mixed β‐adrenoceptor blocker D(+)‐alprenolol all failed to modify the deficit of PA retention by NDO 008 and R(+)‐8‐OH‐DPAT. None of the 5‐HT1A or 5‐HT2A/2C receptor antagonists tested or the β‐blockers altered PA retention by themselves. 5 A 3 day pretreatment procedure (200+100+100 mg kg−1) with the tryptophan hydroxylase inhibitor p‐chlorophenylalanine (PCPA) did not alter PA retention and did not prevent the inhibitory action of the 5‐HT1A agonists, indicating that their effects on PA do not depend on endogenous 5‐HT. 6 The effects of NDO 008 on PA were also studied using a state‐dependent learning paradigm. NDO 008 was found to produce a disruption of PA when given either prior to training or retention or both prior to training and retention but it failed to affect PA retention when given immediately after training. 7 These findings indicate that the deficit of passive avoidance retention induced by the 5‐HT1A agonists is mainly a result of stimulation of postsynaptic 5‐HT1A receptors but not 5‐HT2A receptors. The 5‐HT1A receptor stimulation appears to interfere with learning processes operating at both acquisition and retrieval.

Intraventricular galanin modulates a 5-HT1A receptor-mediated behavioural response in the rat

The present studies have examined whether the neuropeptide galanin can modulate brain serotoninergic (5‐HT) neurotransmission in vivo and, particularly, 5‐HT1A receptor‐mediated transmission. For that purpose, we studied the ability of galanin (given bilaterally into the lateral ventricle, i.c.v.) to modify the impairment of passive avoidance retention induced by the selective 5‐HT1A agonist 8‐hydroxy‐2‐(di‐n‐propyloamino)tetralin (8‐OH‐DPAT) when injected prior to training. This impairment appears to be mainly related to activation of 5‐HT1A receptors in the CNS. Galanin dose‐dependently (significant at 3.0 nmol/rat) attenuated the passive avoidance impairment (examined 24 h after training) induced by the 0.2 mg/kg dose of 8‐OH‐DPAT. This 8‐OH‐DPAT dose produced signs of the 5‐HT syndrome indicating a postsynaptic 5‐HT1A receptor activation. Furthermore, both the impairment of passive avoidance and the 5‐HT syndrome were completely blocked by the 5‐HT1A receptor antagonist WAY 100635 (0.1 mg/kg). Galanin (0.3 or 3.0 nmol) or WAY 100635 (0.1 mg/kg) failed by themselves to affect passive avoidance retention. 8‐OH‐DPAT given at a low dose 0.03 mg/kg, which presumably stimulates somatodendritic 5‐HT1A autoreceptors in vivo, did not alter passive avoidance retention or induce any visually detectable signs of the 5‐HT syndrome. Galanin (0.3 or 3.0 nmol) given i.c.v. in combination with the 0.03 mg/kg dose of 8‐OH‐DPAT, did not modify passive avoidance. The immunohistochemical study of the distribution of i.c.v. administered galanin (10 min after infusion) showed a strong diffuse labelling in the periventricular zone (100–200 μm) of the lateral ventricle. Furthermore, in the dorsal and ventral hippocampus galanin‐immunoreactive nerve cells appeared both in the dentate gyrus and the CA1, CA2 and CA3 layers of the hippocampus. In the septum only endogenous fibres could be seen while in the caudal amygdala also galanin‐immunoreactive nerve cells were visualized far away from the labelled periventricular zone. At the level of the dorsal raphe nucleus a thin periventricular zone of galanin immunoreactivity was seen but no labelling of cells. These results suggest that galanin can modulate postsynaptic 5‐HT1A receptor transmission in vivo in discrete cell populations in forebrain regions such as the dorsal and ventral hippocampus and parts of the amygdala. The indication that galanin administered intracerebroventrically may be taken up in certain populations of nerve terminals in the periventricular zone for retrograde transport suggests that this peptide may also affect intracellular events.

Galanin and learning

A number of studies indicate that galanin (GAL) is a potent modulator of basal acetylcholine release in the rat forebrain e.g. in the cholinergic neurons of the septo-hippocampal projections. Thus, GAL perfused through the microdialysis probe decreased basal acetylcholine release in the ventral hippocampus, while it enhanced acetylcholine release in the dorsal hippocampus. This finding indicates that GAL may act via different mechanisms within the subsystems of the hippocampus. This hypothesis has received support from studies using the Morris swim maze, a learning task dependent on hippocampal mechanisms. GAL (3 nmol/rat) infused into the ventral hippocampus impaired spatial learning acquisition, while it tended to facilitate when injected into the dorsal hippocampus. However, the effects of GAL on acetylcholine release and on spatial learning, which are due to activation of GAL-receptors, appear to be indirectly mediated possibly via noradrenaline transmission. GAL is also a potent inhibitor of mesencephalic 5-HT neurotransmission in vivo. These findings are discussed in relation to the role of acetylcholine and serotonin in cognition.

Prolonged effects of intraventricular galanin on a 5-hydroxytryptamine1A receptor mediated function in the rat

Galanin (3 nmol/rat), 2 h after its intracerebroventricular (i.c.v.) administration to male rats, attenuated the passive avoidance (PA) retention deficit induced by the 5-hydroxytryptamine (HT)(1A) receptor agonist 8-hydroxy-2-(di-N-propylamino)tetraline (8-OH-DPAT) (0.2 mg/kg) The reduction in the postjunctional 5-HT(1A) receptor-mediated response after i.c.v. galanin was not associated with changes in the mRNA levels and agonist binding properties of cortical limbic 5-HT(1A) receptors, believed to be the target receptors mediating the PA deficit caused by 8-OH-DPAT. These results suggest that acute increases of galanin transmission in vivo even after 2 h can counteract limbic 5-HT(1A) receptor-mediated responses of relevance for affective disorders without significantly affecting gene expression and binding characteristics of cortical limbic 5-HT(1A) receptors.

GABA(A) receptor activation in the CA1 area of the dorsal hippocampus impairs consolidation of conditioned contextual fear in C57BL/6J mice

Local infusion of the GABA(A) receptor agonist muscimol is used for reversible inactivation of septohippocampal brain structures associated with cognitive functions. However, information on the effective duration, affected processes and site(s) of action of muscimol in the hippocampus is lacking. Therefore, the dose- and time-dependent effects of bilateral dorsohippocampal infusion of muscimol (0.01-2.0 μg/mouse) below the CA1 area were examined on processing of fear memory in male C57BL/6J mice. Infusion of muscimol 15 min-6 h but not 9 h or 24 h before training impaired conditioned context-dependent fear tested 24 h or 48 h after training. Post-training infusion of muscimol also impaired context-dependent fear when applied either 4 h or 6 h after training, although with lower efficacy. Muscimol was ineffective when administered immediately, 1 h or 24 h after training. Infusion of muscimol 15 min before training impaired context-dependent fear 4-6 h after training indicating preserved short-term but impaired long-term memory. Regardless of infusion time and dose, muscimol had no effect on tone-dependent (cued) fear memory. The impairment by the fluorescently-labeled muscimol-bodipy (5.3 μg/mouse) were similar to those of an equimolar dose of muscimol (1 μg/mouse). The distribution profile after local infusion indicated that muscimol-bodipy (5.3 μg/mouse) was confined to the CA1 area of the dorsal hippocampus. These results demonstrated that GABA(A) receptor activation in the CA1 area of the dorsal hippocampus causes a long-term memory impairment of conditioned context-dependent fear mediated by a long-lasting (≥6 h) muscimol action most likely affecting consolidation processes.