The cognition-enhancing activity of E1R, a novel positive allosteric modulator of sigma-1 receptors.

BACKGROUND AND PURPOSE: Here, we describe the in vitro and in vivo effects of (4R,5S)-2-(5-methyl-2-oxo-4-phenyl-pyrrolidin-1-yl)-acetamide (E1R), a novel positive allosteric modulator of sigma-1 receptors. EXPERIMENTAL APPROACH: E1R was tested for sigma receptor binding activity in a [³H](+)-pentazocine assay, in bradykinin (BK)-induced intracellular Ca²âº concentration ([Ca²âº](i)) assays and in an electrically stimulated rat vas deferens model. E1R's effects on cognitive function were tested using passive avoidance (PA) and Y-maze tests in mice. A selective sigma-1 receptor antagonist (NE-100), was used to study the involvement of the sigma-1 receptor in the effects of E1R. The open-field test was used to detect the effects of E1R on locomotion. KEY RESULTS: Pretreatment with E1R enhanced the selective sigma-1 receptor agonist PRE-084's stimulating effect during a model study employing electrically stimulated rat vasa deferentia and an assay measuring the BK-induced [Ca²âº](i) increase. Pretreatment with E1R facilitated PA retention in a dose-related manner. Furthermore, E1R alleviated the scopolamine-induced cognitive impairment during the PA and Y-maze tests in mice. The in vivo and in vitro effects of E1R were blocked by treatment with the selective sigma-1 receptor antagonist NE-100. E1R did not affect locomotor activity. CONCLUSION AND IMPLICATIONS: E1R is a novel 4,5-disubstituted derivative of piracetam that enhances cognition and demonstrates efficacy against scopolamine-induced cholinergic dysfunction in mice. These effects are attributed to its positive modulatory action on the sigma-1 receptor and this activity may be relevant when developing new drugs for treating cognitive symptoms related to neurodegenerative diseases.

Effects of Hypericum perforatum (St. John's wort) on passive avoidance in the rat: evaluation of potential neurochemical mechanisms underlying its antidepressant activity.

Along with traditional pharmacotherapies, extracts of Hypericum perforatum L. (St. John's wort) are used in the treatment of mild to moderately severe depression. Hypericum is a nonspecific inhibitor of the neuronal uptake of monoamines (serotonin, 5-HT; noradrenaline, NA; dopamine, DA) as well as GABA and glutamate. Hypericum extracts have been shown to be active in several different "animal models for antidepressant drugs". As one of a large number of chemical constituents, the phoroglucinol derivative hyperforin might be an important "antidepressant component" of hypericum. However, the exact role of neurochemical mechanisms underlying in vivo actions of hypericum and hyperforin are not well defined. In the present study, we compared the effects of hypericum, hyperforin and hyperforin-free hypericum and the three conventional antidepressants paroxetine, imipramine and desipramine using the passive avoidance (PA) task in the rat. The 5-HT-releasing compound p-chloroamphetamine (PCA), which operates through the 5-HT neuronal transporter, was used to reveal the potential in vivo effects on 5-HT uptake mechanisms. To examine the ability of the test-compounds to enhance noradrenaline (NA) transmission in vivo, subeffective doses of scopolamine were used. Taken together, our results suggest that (1) hypericum given at high doses can probably affect the neuronal 5-HT uptake mechanisms in a manner more reminiscent of TCAs than SSRIs; (2) similar to TCAs and SSRIs, hypericum and hyperforin are active in the scopolamine test. Hyperforin appears to play a major role in the action of hypericum in this model. Both 5-HT and NA might concomitantly contribute to the effects of different antidepressants in the "low-dose scopolamine" model; (3) hypericum might enhance both 5-HT and NA transmission in forebrain limbic brain circuits important for mood control, which could underly its antidepressant effects. However, the relative contribution of different constituents and exact mechanisms of action require further evaluation.

Prolonged effects of intraventricular galanin on a 5-hydroxytryptamine(1A) 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.

Involvement of the 5-HT1A receptors in classical fear conditioning in C57BL/6J mice.

The present study examined the involvement of the 5-HT(1A) receptors in classical fear conditioning using the 5-HT(1A) agonist 8-hydroxy-2-(di-n-propyloamino)tetralin hydrobromide (8-OH-DPAT) and the selective "silent" 5-HT(1A) receptor antagonist (N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclo- hexane carboxamide trihydrochloride (WAY 100635). The drugs were administered both subcutaneously and bilaterally into the dorsal hippocampus of male C57BL/6J mice. The training was performed in a single trial in which a tone was followed by a footshock. The retention of context- and tone-dependent fear was examined in separate tests conducted either 1 or 24 hr after training. Subcutaneous 8-OH-DPAT (0.1-1.0 mg/kg), when injected before but not after training, caused a dose-dependent impairment of contextual fear in both 1 and 24 hr tests, whereas tone-dependent fear was less affected. Pretraining intrahippocampal injections of 5.0 microg but not 1.0 microg 8-OH-DPAT caused a severe deficit in contextual fear when tested 24 hr after training. When injected both subcutaneously and intrahippocampally, 8-OH-DPAT induced the 5-HT syndrome, indicative of postsynaptic 5-HT(1A) receptor activation at the dose ranges that impaired fear conditioning. However, the behavioral changes induced by 8-OH-DPAT at the time of training could not account for inhibitory effects of 8-OH-DPAT on fear conditioning. Neither subcutaneous (0.03 mg/kg) nor intrahippocampal (0.5 microg per mouse) WAY 100635 altered context- or tone-dependent fear. However, subcutaneous WAY 100635 blocked both the 5-HT syndrome and the impairment of fear conditioning induced by subcutaneous or intrahippocampal 8-OH-DPAT. In contrast, intrahippocampal WAY 100635 blocked the impairment caused by intrahippocampal but not subcutaneous 8-OH-DPAT, indicating the involvement of extrahippocampal 5-HT(1A) receptors in fear conditioning. It is concluded that the deficits in fear conditioning induced by 8-OH-DPAT are a result of postsynaptic 5-HT(1A) receptor activation that interferes with learning processes operating at acquisition but not consolidation. Furthermore, the dorsohippocampal 5-HT(1A) receptors play an important but not exclusive role in the limbic circuitry subserving contextual fear conditioning.

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 Porsolt's 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.

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.

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.

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 s.c.) 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.

Animal models for studying serotonin (5-HT) receptor subtypes: relationship to 5-HT system pathologies.

Despite more than 30 years of intensive research, the specific role of serotonin (5-HT) receptor subtypes in animal models of "anxiety" still remains unclear. The present study focused on the particular role of 5-HT1A receptor subtype in aversive learning, i.e., the passive avoidance (PA) task in the rat. Taken together, the data strongly suggest that: (1) 5-HT1A receptor but not 5-HT2A receptors play a crucial role in PA; (2) the postsynaptic but not presynaptic 5-HT1A receptors are mainly involved in the regulation of PA; (3) 5-HT1A receptors appear to be directly involved both in acquisition and retrieval but not in the consolidation of PA; and (4) besides the "prototypical" 5-HT1A receptor subtype, an additional and yet-unidentified 5-HT receptor subtype seems to play an important modulatory role in PA.

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/beta-adrenoceptor antagonist L(-)-alprenolol. In contrast, the mixed 5-HT2A/2C antagonists ketanserin and pirenperone were found to be ineffective. Moreover, the beta2-adrenoceptor antagonist ICI 118551, the beta-antagonist metoprolol as well as the mixed beta-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 beta-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 microm) 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.

"Atypical" neuromodulatory profile of glutapyrone, a representative of a novel 'class' of amino acid-containing dipeptide-mimicking 1,4-dihydropyridine (DHP) compounds: in vitro and in vivo studies.

Glutapyrone, a disodium salt of 2-(2,6-dimethyl-3,5-diethoxycarbonyl-1,4-dihydropyridine-4-carboxamido)- glutaric acid, is a representative of a novel 'class' of amino acid-containing 1,4-dihydropyridine (DHP) compounds developed at the Latvian Institute of Organic Synthesis, Riga, Latvia. Conceptually, the glutapyrone molecule can be regarded as a dipeptide-mimicking structure formed by the "free" amino acid (glutamate) moiety and "crypto" (built into the DHP cycle) amino acid ("GABA") elements. Both of these amino acids are joined by the peptide bond. This compound unlike classical DHPs lacks calcium antagonistic or agonistic properties. Our previous studies revealed a profound and long-term anticonvulsant, stress-protective and neurodeficit-preventive activities of glutapyrone. In view of structural properties the role of glutamatergic mechanisms in the mediation of central effects of glutapyrone was considered. In the present study glutapyrone at the concentration range of 1 microM(-1) mM failed to effect both NMDA ([3H]TCP) and non-NMDA ([3H]KA and [3H]AMPA) receptor ligand binding in the rat cortical membranes in vitro. The compound markedly enhanced motor hyperactivity induced by the NMDA antagonist PCP and the dopamine releasing compound D-amphetamine in the rats. Glutapyrone displayed activity in a variety of animal models relevant for affective/depressive disorders in humans i.e. reserpine-induced ptosis and hypothermia, forced swimming test and open field test. These data indicate that the unusually "broad" pharmacological spectrum of glutapyrone might involve concomitant actions on multiple neurotransmitter systems, particularly, GABA-ergic and the catecholamines. It is discussed whether these functional properties are secondary to action on intracellular events, predominantly, G protein-related since glutapyrone appears to lack direct interactions with a number of receptors including ionotropic glutamate and GABA(A)/Bzd receptors.

Galanin modulates 5-hydroxytryptamine functions. Focus on galanin and galanin fragment/5-hydroxytryptamine1A receptor interactions in the brain.

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 anti-depressant 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.

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.