Serotonergic enhancement of circadian responses to light: role of the raphe and intergeniculate leaflet

Light serves as the primary stimulus that synchronizes the circadian clock in the suprachiasmatic nucleus (SCN) to the external day/night cycle. Appropriately timed light exposure can reset the phase of the circadian clock. Some serotonergic drugs that bind to the serotonin 1A receptor can enhance phase shifts to light. The mechanism by which this potentiation occurs is not well understood. In this study, we examined where one of these drugs, 8‐[2‐[4‐(2‐methoxyphenyl)‐1‐piperazinyl]ethyl]‐8‐azaspiro[4.5]decane‐7,9‐dione dihydrochloride (BMY7378), might be working in the hamster brain. Systemic (5 mg/kg), intra‐dorsal raphe and intra‐median raphe (both 15.6 nmol in 0.5 μL), but not intra‐SCN (7.8 nmol or 15.6 nmol in 0.5 μL) injections of BMY7378 significantly potentiated phase shifts to light. Potentiation of photic shifts persisted when serotonergic innervation of the SCN was lesioned with infusions of the serotonin neurotoxin 5,7‐dihydroxytryptamine into the SCN. Light‐induced c‐Fos expression in the rostral and caudal intergeniculate leaflet (IGL) was attenuated with systemic BMY7378, suggesting that the IGL may be involved in this response. Both complete IGL lesions and depletion of serotonergic innervation of the IGL prevented systemic BMY7378 from potentiating photic phase shifts. Together, these findings suggest that the mechanism by which BMY7378 enhances photic responses is by changing the activity of the raphe nuclei to influence how the IGL responds to light, which subsequently influences the SCN as one of its downstream targets. Identification of the network that underlies this potentiation could lead to the development of useful therapeutic interventions for treating sleep and circadian disorders.     

The serotonergic anxiolytic buspirone attenuates circadian responses to light

Serotonergic drugs modify circadian responses to light, with agonists attenuating and some partial agonists or antagonists potentiating photic phase shifts. The anxiolytic buspirone is a 5‐HT_1A receptor partial agonist. Given that buspirone is used therapeutically to manage generalised anxiety disorder, it would be useful to understand if and how this drug may modify circadian responses to light, not only to help manage side effects, but also to examine its potential use as a chronobiotic. Here we examined behavioral and molecular responses to phase‐shifting light in mice and hamsters treated with buspirone. Phase advances to late subjective night light pulses in hamsters and wildtype mice were significantly attenuated by buspirone. 5‐HT_1A receptor knockout mice exhibited potentiated photic phase shifts when pretreated with buspirone. In wildtype mice, the attenuated phase shifts were accompanied by increased cFos expression in the suprachiasmatic nucleus, whereas potentiated phase shifts in knockouts were accompanied by increased phosphorylation of extracellular signal‐regulated kinase (ERK) and cyclic AMP response element‐binding protein (CREB), and decreased cFos expression. Attenuated photic phase shifts in buspirone‐treated hamsters were accompanied by decreased phosphorylation of ERK and CREB. Chronic buspirone treatment decreased the amplitude of wheel‐running rhythms, lengthened the duration of the active phase and advanced the phase angle of entrainment. Buspirone administration at midday produced non‐photic phase advances in wildtype but not 5‐HT_1A receptor knockout mice. These findings suggest that buspirone affected the circadian system in a manner similar to the 5‐HT_1A/7 agonist (±)‐8‐Hydroxy‐2‐dipropylaminotetralin hydrobromide, primarily through the 5‐HT_1A receptor, and suggest that therapeutic use of buspirone to manage anxiety may impact circadian function.     

Activation of 5‐HT receptors inhibits GABAergic transmission by pre‐and post‐synaptic mechanisms in layer II/III of the juvenile rat auditory cortex

The specific mechanisms by which serotonin (5‐HT) modulates synaptic transmission in the auditory cortex are still unknown. In this work, we used whole‐cell recordings from layer II/III of pyramidal neurons in rat brain slices to characterize the influence of 5‐HT on inhibitory synaptic activity in the auditory cortex after pharmacological blockade of excitatory glutamatergic transmission. We found that bath application of 5‐HT (5 µM) reduced the frequency and amplitude of both spontaneous and miniature inhibitory postsynaptic currents (IPSCs), reduced the amplitude of evoked IPSCs, and enhanced facilitation of paired pulse ratio (PPR), suggesting presynaptic inhibition. To determine which the serotonin receptors were involved in this effect, we studied the influence of specific 5‐HT receptor agonists and antagonists on ?‐aminobutyric acid (GABA)ergic synaptic transmission. The inhibiting influence of 5‐HT in the GABAergic synaptic activity was mimicked by using the selective agonists of the 5‐HT_1A and 5‐HT_2A receptors, 8(OH)‐DPAT (10 µM) and DOI (10 µM), respectively; and it was prevented by their respective antagonists NAN‐190 (1 µM) and ritanserin (1 μM). Furthermore, the application of the selective agonist of 5‐HT_1A receptors, 8‐(OH)‐DPAT (10 µM), produced PPR facilitation, while DOI application (5‐HT_2A agonist) did not change the PPR. Moreover, the 5‐HT_2A agonist reduced the amplitude of the IPSCs evoked by application of the selective GABA agonist, muscimol. These results suggest a presynaptic and postsynaptic reduction of GABAergic transmission mediated by 5‐HT_1A and 5‐HT_2A serotonergic receptors, respectively. Synapse 69:115–127, 2015. © 2014 Wiley Periodicals, Inc.     

Brain 5-HT(2A) receptors in MPTP monkeys and levodopa-induced dyskinesias

Levodopa‐induced dyskinesias (LIDs) are abnormal involuntary movements induced by the chronic use of levodopa (l ‐Dopa) limiting the quality of life of Parkinson’s disease (PD) patients. We evaluated changes of the serotonin 5‐HT_2A receptors in control monkeys, in 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐lesioned monkeys and in l ‐Dopa‐treated MPTP monkeys, without or with adjunct treatments to inhibit the expression of LID: CI‐1041, a selective NR1A/2B subunit antagonist of glutamate N‐methyl‐d ‐aspartic acid (NMDA) receptor, or Cabergoline, a long‐acting dopamine D₂ receptor agonist. All treatments were administered for 1 month and animals were killed 24 h after the last dose of l ‐Dopa. Striatal concentrations of serotonin were decreased in all MPTP monkeys investigated, as measured by high‐performance liquid chromatography. [³H]Ketanserin‐specific binding to 5‐HT_2A receptors was measured by autoradiography. l ‐Dopa treatment that induced dyskinesias increased 5‐HT_2A receptor‐specific binding in the caudate nucleus and the anterior cingulate gyrus (AcgG) compared with control monkeys. Moreover, [³H]Ketanserin‐specific binding was increased in the dorsomedial caudate nucleus in l ‐Dopa‐treated MPTP monkeys compared with saline‐treated MPTP monkeys. Nondyskinetic monkeys treated with CI‐1041 or Cabergoline showed low 5‐HT_2A‐specific binding in the posterior dorsomedial caudate nucleus and the anterior AcgG compared with dyskinetic monkeys. No significant difference in 5‐HT_2A receptor binding was observed in any brain regions examined in saline‐treated MPTP monkeys compared with control monkeys. These results confirm the involvement of serotonergic pathways and the glutamate/serotonin interactions in LID. They also support targeting 5‐HT_2A receptors as a potential treatment for LID.     

Serotonin‐2C receptor agonists decrease potassium‐stimulated GABA release in the nucleus accumbens

The serotonin 5‐HT_2C receptor has shown promise in vivo as a pharmacotherapeutic target for alcoholism. For example, recently, a novel 4‐phenyl‐2‐N,N‐dimethylaminotetralin (PAT) drug candidate, that demonstrates 5‐HT_2C receptor agonist activity together with 5‐HT_2A/2B receptor inverse agonist activity, was shown to reduce operant responding for ethanol after peripheral administration to rats. Previous studies have shown that the 5‐HT_2C receptor is found throughout the mesoaccumbens pathway and that 5‐HT_2C receptor agonism causes activation of ventral tegmental area (VTA) GABA neurons. It is unknown what effect 5‐HT_2C receptor modulation has on GABA release in the nucleus accumbens core (NAcc). To this end, microdialysis coupled to capillary electrophoresis with laser‐induced fluorescence was used to quantify extracellular neurotransmitter concentrations in the NAcc under basal and after potassium stimulation conditions, in response to PAT analogs and other 5‐HT_2C receptor modulators administered by reverse dialysis to rats. 5‐HT_2C receptor agonists specifically attenuated stimulated GABA release in the NAcc while 5‐HT_2C antagonists or inverse agonists had no effect. Agents with activity at 5‐HT_2A receptors had no effect on GABA release. Thus, in contrast to results reported for the VTA, current results suggest 5‐HT_2C receptor agonists decrease stimulated GABA release in the NAcc, and provide a possible mechanism of action for 5HT_2C‐mediated negative modulation of ethanol self‐administration. Synapse 69:78–85, 2015. © 2014 Wiley Periodicals, Inc.     

Triptans attenuate circadian responses to light

Daily exposure to light synchronizes the circadian clock, located in the suprachiasmatic nucleus (SCN), to external day/night cycles. These responses to light can be modified by serotonergic drugs, such as serotonin 5HT_1B receptor agonists. Triptans are specific 5HT_1B agonists prescribed to treat migraines. Here, we examined the effects of two triptans (zolmitriptan and sumatriptan) on photic phase resetting in Syrian hamsters. Pre‐treatment with intra‐SCN sumatriptan significantly attenuates, and at higher doses completely blocks, phase advances to light during the late night. Pre‐treatment with systemic zolmitriptan significantly attenuates both light‐induced phase advances and phase delays. Neither of these drugs, nor their vehicles, causes phase shifts on their own. Pre‐treatment with zolmitriptan also significantly reduces the expression of light‐induced c‐fos in the SCN. Neither zolmitriptan nor vehicle alone induces significant c‐fos expression in the SCN. Finally, pre‐treatment with zolmitriptan does not attenuate phase shifts to intra‐SCN N‐methyl‐d ‐aspartate injections, indicating that the mechanism of action for zolmitriptan is likely to be through activation of presynaptic 5HT_1B receptors on retinal terminals, thereby decreasing light‐induced neurotransmitter release. As triptans are commercially available medications, there is potential for their use in blocking unwanted photic phase shifting during shift‐work or jet‐lag. Additionally, triptans may also affect the circadian clock in patients receiving them regularly for migraines. Finally, our results may hint at the mechanism by which triptans can alleviate the photophobia that frequently accompanies migraines, namely by activating 5HT_1B receptors on retinal terminals elsewhere in the brain, and thereby diminishing visually‐evoked neurotransmitter signalling in those areas.     

Cannabinoid receptor agonists upregulate and enhance serotonin 2A (5‐HT2A) receptor activity via ERK1/2 signaling

Recent behavioral studies suggest that nonselective agonists of cannabinoid receptors may regulate serotonin 2A (5‐HT_2A) receptor neurotransmission. Two cannabinoids receptors are found in brain, CB1 and CB2 receptors, but the molecular mechanism by which cannabinoid receptors would regulate 5‐HT_2A receptor neurotransmission remains unknown. Interestingly, we have recently found that certain cannabinoid receptor agonists can specifically upregulate 5‐HT_2A receptors. Here, we present experimental evidence that rats treated with a nonselective cannabinoid receptor agonist (CP 55,940, 50 µg/kg, 7 days) showed increases in 5‐HT_2A receptor protein levels, 5‐HT_2A receptor mRNA levels, and 5‐HT_2A receptor‐mediated phospholipase C beta (PLCβ) activity in prefrontal cortex (PFCx). Similar effects were found in neuronal cultured cells treated with CP 55,940 but these effects were prevented by selective CB2, but not selective CB1, receptor antagonists. CB2 receptors couple to the extracellular kinase (ERK) signaling pathway by Gα_i/o class of G‐proteins. Noteworthy, GP 1a (selective CB2 receptor agonist) produced a strong upregulation of 5‐HT_2A receptor mRNA and protein, an effect that was prevented by selective CB2 receptor antagonists and by an ERK1/2 inhibitor, PD 198306. In summary, our results identified a strong cannabinoid‐induced upregulation of 5‐HT_2A receptor signaling in rat PFCx. Our cultured cell studies suggest that selective CB2 receptor agonists upregulate 5‐HT_2A receptor signaling by activation of the ERK1/2 signaling pathway. Activity of cortical 5‐HT_2A receptors has been associated with several physiological functions and neuropsychiatric disorders such as stress response, anxiety and depression, and schizophrenia. Therefore, these results may provide a molecular mechanism by which activation of cannabinoid receptors might be relevant to the pathophysiology of some cognitive and mood disorders in humans. Synapse, 2013. © 2012 Wiley Periodicals, Inc.     

Effect of a hallucinogenic serotonin 5‐HT2A receptor agonist on visually guided,hippocampal‐dependent spatial cognition in C57BL/6J mice

By acting on serotonin 5‐HT_2A receptors (5‐HT_2ARs), serotonergic psychedelic drugs induce perceptual and visual hallucinations by increasing neuronal excitability and altering visual‐evoked neuronal responses. The present study was designed to examine whether the perceptual alterations induced by a serotonergic psychedelic drug would affect the integrity of hippocampal‐dependent, visually guided spatial cognition. phenylalkylamine hallucinogen TCB‐2 is a selective agonist of 5‐HT_2ARs. Mice received TCB‐2 (1.0 mg kg⁻¹, i.p.), and spatial behaviors and hippocampal electrophysiological responses were measured with water maze tasks and in vivo single‐unit recording, respectively. TCB‐2 did not affect visual cue approach behavior in the visible platform water maze, but increased the latency of trained mice to initiate goal‐directed swimming during a probe test in the hidden platform Morris water maze, which could be prevented by 5‐HT_2AR antagonist MDL 11,939. Interestingly, TCB‐2 did not affect the efficiency of the swim path or the proper use of distal visual cues during the probe test. Hippocampal place cell activity is considered to represent spatial and context‐specific episodic memory. Systemic TCB‐2 did not affect previously established place fields of CA1 neurons in mice exploring a familiar environment, or the remapping of place cells when the mice explored a novel environment. However, TCB‐2 impaired the long‐term stability of place fields for the novel environment initially encoded under the influence of TCB‐2, which could be prevented by 5‐HT_2AR antagonist MDL 11,939. Our data indicate that hallucinogenic 5‐HT_2AR agonist delays the initiation of spatial search behavior, but does not impair the use of visual cues to guide goal‐directed spatial behavior. Moreover, activation of 5‐HT_2ARs does not impair the coding and retrieval of spatial information, but impairs the long‐term stability of new formed place fields of CA1 neurons. © 2017 Wiley Periodicals, Inc.     

Differential effects of serotonin (5‐HT)2 receptor‐targeting ligands on locomotor responses to nicotine‐repeated treatment

We verified the hypothesis that serotonin (5‐HT)₂ receptors control the locomotor effects of nicotine (0.4 mg kg⁻¹) in rats by using the 5‐HT_2A receptor antagonist M100907, the preferential 5‐HT_2A receptor agonist DOI, the 5‐HT_2C receptor antagonist SB 242084, and the 5‐HT_2C receptor agonists Ro 60‐0175 and WAY 163909. Repeated pairings of a test environment with nicotine for 5 days, on Day 10 significantly augmented the locomotor activity following nicotine administration. Of the investigated 5‐HT₂ receptor ligands, M100907 (2 mg kg⁻¹) or DOI (1 mg kg⁻¹) administered during the first 5 days in combination with nicotine attenuated or enhanced, respectively, the development of nicotine sensitization. Given acutely on Day 10, M100907 (2 mg kg⁻¹), Ro 60‐0175 (1 mg kg⁻¹), and WAY 163909 (1.5 mg kg⁻¹) decreased the expression of nicotine sensitization. In another set of experiments, where the nicotine challenge test was performed on Day 15 in animals treated repeatedly (Days: 1–5, 10) with nicotine, none of 5‐HT₂ receptor ligands administered during the second withdrawal period (Days: 11–14) to nicotine‐treated rats altered the sensitizing effect of nicotine given on Day 15. Our data indicate that 5‐HT_2A receptors (but not 5‐HT_2C receptors) play a permissive role in the sensitizing effects of nicotine, while stimulation of 5‐HT_2A receptors enhances the development of nicotine sensitization and activation of 5‐HT_2C receptors is essential for the expression of nicotine sensitization. Repeated treatment with the 5‐HT₂ receptor ligands within the second nicotine withdrawal does not inhibit previously established sensitization. Synapse 64:511–519, 2010. © 2010 Wiley‐Liss, Inc.     

Hippocampal activation of 5‐HT1B receptors and BDNF production by vagus nerve stimulation in rats under chronic restraint stress

A growing body of evidence shows that the electrical stimulation of the vagus nerve can improve mental illness including depression. Here, we investigated whether the vagus nerve stimulation (VNS) is involved in regulating the responsiveness of hippocampal neurons in rats under chronic restraint stress (CRS). c‐Fos protein signals were detected 2 hr after VNS in 5‐HT_1A receptor‐positive neurons in the dorsal raphe nucleus (DRN) as well as in the nucleus tractus solitarius (NTS). Chronic VNS was performed on a daily basis for 2 weeks using an implanted microelectrode in rats that had undergone CRS for 2 weeks. We found that the levels of both 5‐HT_1B receptors and phospho‐Erk1/2 were decreased in parallel in the hippocampal neurons of CRS animals and then increased to the baseline levels by chronic VNS. Hippocampal induction of 5‐HT_1B receptors and phospho‐Erk1/2 by VNS was diminished after the injection of 5,7‐dihydroxytryptamine (5,7‐DHT), a neurotoxin of serotonergic neurons, into the DRN. Hippocampal production of brain‐derived neurotrophic factor (BDNF) was also upregulated by VNS, but the treatment of 5,7‐DHT abrogated the effects of VNS on BDNF induction. VNS in CRS animals improved the behavioral scores in forced swimming test (FST) compared to sham‐stimulated control. Our results suggest that VNS‐mediated serotonergic input via 5‐HT_1B receptors into the hippocampal neurons may activate BDNF pathway and improve depressive‐like behaviors in CRS animals.     

Relations between cortical thickness,serotonin 1A receptor binding,and structural connectivity: A multimodal imaging study

Serotonin 1A (5‐HT_1A) receptors play a direct role in neuronal development, cell proliferation, and dendritic branching. We hypothesized that variability in 5‐HT_1A binding can affect cortical thickness, and may account for a subtype of major depressive disorder (MDD) in which both are altered. To evaluate this, we measured cortical thickness from structural magnetic resonance imaging (MRI) and 5‐HT_1A binding by positron emission tomography (PET) in an exploratory study. To examine a range of 5‐HT_1A binding and cortical thickness values, we recruited 25 healthy controls and 19 patients with MDD. We hypothesized increased 5‐HT_1A binding in the raphe nucleus (RN) would be negatively associated with cortical thickness due to reduced serotonergic transmission. Contrary to our hypothesis, raphe 5‐HT_1A binding was positively correlated with cortical thickness in right posterior cingulate cortex (PCC), a region implicated in the default mode network. Cortical thickness was also positively correlated with 5‐HT_1A in each cortical region. We further hypothesized that the strength of 5‐HT_1A‐cortical thickness correlation depends on the number of axons between the raphe nucleus and each region. To explore this we related 5‐HT_1A–cortical thickness correlation coefficients to the number of tracts connecting that region and the raphe, as measured by diffusion tensor imaging (DTI) in an independent sample. The 5‐HT_1A–cortical thickness association correlated significantly with the number of tracts to each region, supporting our hypothesis. We posit a defect in the raphe may affect the PCC within the default mode network in MDD through serotonergic fibers, resulting in increased ruminative processing.     

Serotonin (5‐HT) regulates neurite outgrowth through 5‐HT1A and 5‐HT7 receptors in cultured hippocampal neurons

Serotonin (5‐HT) production and expression of 5‐HT receptors (5‐HTRs) occur early during prenatal development. Recent evidence suggests that, in addition to its classical role as a neurotransmitter, 5‐HT regulates neuronal connectivity during mammalian development by modulating cell migration and neuronal cytoarchitecture. Given the variety of 5‐HTRs, researchers have had difficulty clarifying the specific role of each receptor subtype in brain development. Signalling mediated by the G‐protein‐coupled 5‐HT_1AR and 5‐HT₇R, however, has been associated with neuronal plasticity. Thus, we hypothesized that 5‐HT promotes neurite outgrowth through 5‐HT_1AR and 5‐HT₇R. The involvement of 5‐HT_1AR and 5‐HT₇R in the morphology of rat hippocampal neurons was evaluated by treating primary cultures at 2 days in vitro with 5‐HT and specific antagonists for 5‐HT_1AR and 5‐HT₇R (WAY‐100635 and SB269970, respectively). The stimulation of hippocampal neurons with 100 nM 5‐HT for 24 hr produced no effect on either the number or the length of primary neurites. Nonetheless, after 5HT₇R was blocked, the addition of 5‐HT increased the number of primary neurites, suggesting that 5HT₇R could inhibit neuritogenesis. In contrast, 5‐HT induced secondary neurite outgrowth, an effect inhibited by 1 μM WAY‐100635 or SB269970. These results suggest that both serotonergic receptors participate in secondary neurite outgrowth. We conclude that 5‐HT_1AR and 5‐HT₇R regulate neuronal morphology in primary hippocampal cultures by promoting secondary neurite outgrowth. © 2014 Wiley Periodicals, Inc.     

5-HT1A agonists induce hippocampal theta activity in freely moving cats: role of presynaptic 5-HT1A receptors

Electrical activity in the dorsal hippocampus was recorded in freely moving cats in response to intravenous administration of 5-HT_1A agonist and antagonist drugs. Administration of low doses of the selective 5-HT_1A agonists 8-OH-DPAT (5–20 μg/kg) and ipsapirone (20–100 μg/kg) produced rhythmic slow activity (theta) in the hippocampal EEG within 30 s. Similar effects were observed with BMY 7378 (20 and 100 μg/kg), which acts as an agonist at presynaptic (somatodendritic) 5-HT_1A receptors and as an antagonist at postsynaptic 5-HT_1A receptors. Power spectral analyses showed that all three compounds produced a dose-dependent increase in the EEG power occurring in the theta frequency band (3.5–8.0 Hz) as a proportion of total power from 0.25 to 30.0 Hz (relative theta power). The increase in relative theta power produced by 8-OH-DPAT (20 μg/kg) was greatly attenuated by spiperone (1 mg/kg), a highly effective 5-HT_1A autoreceptor antagonist. Administration of spiperone alone had no significant effect on relative theta power. These results are discussed in relationship to the effects of these drugs on serotonergic neuronal activity. Our results suggest that preferential activation of presynaptic 5-HT_1A receptors, and subsequent inhibition of serotonin neurotransmission, facilitates the appearance of hippocampal theta activity in awake cats.     

Site‐specific effects of gastrin‐releasing peptide in the suprachiasmatic nucleus

The effects of gastrin‐releasing peptide (GRP) on the circadian clock in the suprachiasmatic nucleus (SCN) are dependent on the activation of N‐methyl‐d ‐aspartate (NMDA) receptors in the SCN. In this study, the interaction between GRP, glutamate and serotonin in the regulation of circadian phase in Syrian hamsters was evaluated. Microinjection of GRP into the third ventricle induced c‐fos and p‐ERK expression throughout the SCN. Coadministration of an NMDA antagonist or 8‐hydroxy‐2‐di‐n‐propylamino‐tetralin [a serotonin (5‐HT)_1A,7 agonist, DPAT] with GRP limited c‐fos expression in the SCN to a region dorsal to GRP cell bodies. Similar to the effects of NMDA antagonists, DPAT attenuated GRP‐induced phase shifts in the early night, suggesting that the actions of serotonin on the photic phase shifting mechanism occur downstream from retinorecipient cells. c‐fos and p‐ERK immunoreactivity in the supraoptic (SON) and paraventricular hypothalamic nuclei also increased following ventricular microinjection of GRP. Because of this finding, a second set of experiments was designed to test a potential role for the SON in the regulation of clock function. Syrian hamsters were given microinjections of GRP into the peri‐SON during the early night. GRP‐induced c‐fos activity in the SCN was similar to that following ventricular administration of GRP. GRP or bicuculline (a γ‐aminobutyric acid_A antagonist) administered near the SON during the early night elicited phase delays of circadian activity rhythms. These data suggest that GRP‐induced phase‐resetting is dependent on levels of glutamatergic and serotonergic neurotransmission in the SCN and implicate activity in the SON as a potential regulator of photic signaling in the SCN.     

Acute and chronic effects of citalopram on 5‐HT1A receptor—Labeling by [18F]MPPF and—Coupling to receptors‐G proteins

Selective serotonin reuptake inhibitors take several weeks to produce their maximal therapeutic antidepressant effect. This delay has been attributed to the gradual desensitization of somatodendritic serotonin 5‐HT_1A autoreceptors. We evaluated adaptive changes of 5‐HT_1A receptors after acute and chronic citalopram challenges in rat. Small animal positron emission tomography trial and quantitative ex vivo autoradiography studies using [¹⁸F]MPPF were employed, as well as in vitro 8‐OH‐DPAT‐stimulated [³⁵S]‐GTPγS binding assay. Additionally, 5‐HT_1A receptor knock‐out mice were used to assess the specificity of [¹⁸F]MPPF. Acute treatment with citalopram did not alter [¹⁸F]MPPF binding in dorsal raphe nucleus (DR), frontal cortex, or hippocampus. The absence of [¹⁸F]MPPF binding in the brain of 5‐HT_1A knock‐out mice demonstrates the specificity of MPPF for 5‐HT_1A receptor brain imaging, but the high affinity of [¹⁸F]MPPF compared to 5‐HT suggests that it would only be displaced by dramatic increases in extracellular 5‐HT. Chronic citalopram did not modify 5‐HT_1A receptor density in any of the brain regions studied. In addition, this treatment did not modify 8‐OH‐DPAT‐stimulated [³⁵S]‐GTPγS binding in DR, although a significant increase was observed in frontal cortex and hippocampus. [¹⁸F]MPPF appears to be an efficient radioligand to quantify specifically 5‐HT_1A receptor density in brain imaging. The delayed therapeutic efficacy of citalopram did not appear to be linked to either a downregulation of 5‐HT_1A receptors or to a 5‐HT_1A receptor‐G protein decoupling process in serotonergic neurons, but to increased functional sensitivity of postsynaptic 5‐HT_1A receptors. Synapse 63:106–116, 2009. ©2008 Wiley‐Liss, Inc.     

Serotonin excites hippocampal CA1 GABAergic interneurons at the stratum radiatum‐stratum lacunosum moleculare border

The hippocampus receives robust serotonergic innervation that is thought to control the excitability of both pyramidal cells and GABAergic interneurons. Previous work has addressed serotonergic regulation of pyramidal cells but considerable gaps remain in our understanding of how serotonin regulates different interneuron subclasses. 5‐HT_2A receptors (5‐HT_2ARs) appear to localize predominantly, if not solely, on interneurons in the hippocampus and have been implicated in the regulation of hippocampal function including mnemonic and novelty recognition processes. Interneurons are functionally diverse. Therefore in the current work, we have used a BAC transgenic mouse line expressing EGFP under the control of the 5‐HT_2AR promoter to identify the interneuron subtype(s) regulated by serotonin via 5‐HT_2ARs. We find that EGFP expression in this mouse identifies a group of interneurons that resides predominantly along the border of the stratum radiatum (SR) and stratum lacunosum moleculare (SLM) of the CA1 region. We then show that these cells are depolarized and excited by serotonin acting through 5‐HT_2ARs and appear to belong predominantly to the perforant pathway‐associated and Schaffer collateral/commissural pathway‐associated subtypes. These results indicate that serotonin interneurons expressing 5‐HT_2ARs are localized primarily along the SR‐SLM border of the CA1 region and represent a newly identified target for serotonin regulation in the hippocampus. © 2016 Wiley Periodicals, Inc.     

5‐HT1A and NMDA receptors interact in the rat medial septum and modulate hippocampal‐dependent spatial learning

Cholinergic and GABAergic neurons in the medial septum/vertical limb of the diagonal band of Broca (MS/vDB) projecting to the hippocampus, constitute the septohippocampal projection, which is important for hippocampal‐dependent learning and memory. There is also evidence for an extrinsic as well as an intrinsic glutamatergic network within the MS/vDB. GABAergic and cholinergic septohippocampal neurons express the serotonergic 5‐HT_1A receptor and most likely also glutamatergic NMDA receptors. The aim of the present study was to examine whether septal 5‐HT_1A receptors are important for hippocampal‐dependent long‐term memory and whether these receptors interact with glutamatergic NMDA receptor transmission in a manner important for hippocampal‐dependent spatial memory. Intraseptal infusion of the 5‐HT_1A receptor agonist (R)‐8‐OH‐DPAT (1 or 4 μg/rat) did not affect spatial learning in the water maze task but impaired emotional memory in the passive avoidance task at the higher dose tested (4 μg/rat). While intraseptal administration of (R)‐8‐OH‐DPAT (4 μg) combined with a subthreshold dose of the NMDA receptor antagonist D‐AP5 (1 μg) only marginally affected spatial acquisition, it produced a profound impairment in spatial memory. In conclusion, septal 5‐HT_1A receptors appears to play a more prominent role in emotional than in spatial memory. Importantly, septal 5‐HT_1A and NMDA receptors appear to interact in a manner, which is particularly critical for the expression or retrieval of hippocampal‐dependent long‐term spatial memory. It is proposed that NMDA receptor hypofunction in the septal area may unmask a negative effect of 5‐HT_1A receptor activation on memory, which may be clinically relevant. © 2009 Wiley‐Liss, Inc.     

Juvenile 5HT1B receptor knockout mice exhibit reduced pharmacological sensitivity to 5HT1A receptor activation

Serotonin is an important modulator of anxiety and thus drugs that act on this system have frequently been shown to be either anxiogenic or anxiolytic. In addition serotonin has important trophic functions during early development and disruption of serotonin homeostasis is likely to have long-lasting repercussions in the adult. In the present study we examined the contribution of two serotonin receptor subtypes (5HT_1A and 5HT_1B) to the pathophysiology of anxiety during development. For this, we have studied homozygous knockout mice lacking the 5HT_1B receptor and examined the effect of pharmacological manipulations of 5HT_1A and 5HT_1B receptors on locomotor activity and emission of ultrasonic vocalization (USV) in 7–8 days old mice. As shown before, drug naïve 5HT_1B knockout pups showed reduced USV and were hyperactive, in comparison to wild type controls. The administration of RU24969 (a 5HT_1A/1B agonist) showed a dose-dependent decrease in USV in the wild type and a biphasic effect in the mutants and resulted in dose-dependent increase in activity in the wild type and, to a lesser extent, in the knockouts. The selective 5HT_1A agonist, 8OH-DPAT, dose-dependently blocked vocalization in both genotypes and also increased locomotion. To differentially activate 5HT_1B receptors we first blocked 5HT_1A receptors with WAY100315 and then treated with RU24969. At a high testing temperature, pretreatment with WAY100315 resulted in an anxiogenic effect in wild type pups but not in the knockouts. In agreement with our findings that 5HT_1B knockout mice were in general less sensitive to 5HT_1A activation, 5HT_1A receptor binding was reduced in the knockouts in comparison to controls. Finally, treatment with diazepam dose-dependently decreased USVs in both group with the knockouts showing enhanced sensitivity to this drug. Our results show that important adaptations to a disturbance of serotonin homeostasis occur during the first week of life within the serotonergic system. The observed decreased in sensitivity of 5HT_1B knockout mice to 5HT_1A and increased to GABA_A manipulations are discussed within the context of serotonergic plasticity during development and the implication for clinical treatment of anxiety in genetically predisposed individuals.     

Interactions of serotonin (5‐HT)2 receptor‐targeting ligands and nicotine: Locomotor activity studies in rats

Male Wistar rats were used to verify the hypothesis that serotonin (5‐HT)_2A or 5‐HT_2C receptors may control the locomotor effects evoked by nicotine (0.4mg/kg). The 5‐HT_2A receptor antagonist (M100,907), the 5‐HT_2A receptor agonist (DOI), the 5‐HT_2C receptor antagonist (SB 242,084), and the 5‐HT_2C receptor agonists (Ro 60‐0175 and WAY 163,909) were used. M100,907 (0.5–2mg/kg) did not alter, while DOI (1 mg/kg) enhanced the nicotine‐induced hyperlocomotion. The effect of DOI was antagonized by M100,907 (1 mg/kg). SB 242,084 (0.25–1 mg/kg) augmented, while Ro 60‐0175 (1 and 3 mg/kg) and WAY 163,909 (1.5 mg/kg) decreased the overall effect of acute nicotine; effects of Ro 60‐0175 and WAY 163,909 were attenuated by SB 242,084 (0.125 mg/kg). In another set of experiments, M100,907 (2 mg/kg) on Day 10 attenuated, while DOI (0.1–1 mg/kg) enhanced the nicotine‐evoked conditioned hyperlocomotion in rats repeatedly (Days 1–5) treated with nicotine in experimental chambers. SB 242,084 (0.125 or 1 mg/kg) did not change, while Ro 60‐0175 (1 mg/kg) or WAY 163,909 (1.5 mg/kg) decreased the expression of nicotine‐induced conditioned hyperactivity. Only DOI (0.3 and 1 mg/kg) and SB 242,084 (1 mg/kg) enhanced the basal locomotion. The present data indicate that 5‐HT_2A receptors are significant for the expression of nicotine‐evoked conditioned hyperactivity. Conversely, 5‐HT_2C receptors play a pivotal role in the acute effects of nicotine. Pharmacological stimulation of 5‐HT_2A receptors enhances the conditioned hyperlocomotion, while activation of 5‐HT_2C receptors decreases both the response to acute nicotine and conditioned hyperactivity. Synapse 63:653–661, 2009. © 2009 Wiley‐Liss, Inc.     

An immunocapture/scintillation proximity analysis of Gαq/11 activation by native serotonin (5‐HT)2A receptors in rat cortex: Blockade by clozapine and mirtazapine

Though transduction mechanisms recruited by heterologously expressed 5‐HT_2A receptors have been extensively studied, their interaction with specific subtypes of G‐protein remains to be directly evaluated in cerebral tissue. Herein, as shown by an immunocapture/scintillation proximity analysis, 5‐HT, the prototypical 5‐HT_2A agonist, DOI, and Ro60,0175 all enhanced [³⁵S]GTPγS binding to Gαq/11 in rat cortex with pEC₅₀ values of 6.22, 7.24 and 6.35, respectively. No activation of Go or Gs/olf was seen at equivalent concentrations of DOI. Stimulation of Gαq/11 by 5‐HT (30 μM) and DOI (30 μM) was abolished by the selective 5‐HT_2A vs. 5‐HT_2C/5‐HT_2B antagonists, ketanserin (pK_B values of 9.11 and 8.88, respectively) and MDL100,907 (9.82 and 9.68). By contrast, 5‐HT‐induced [³⁵S]GTPγS binding to Gαq/11 was only weakly inhibited by the preferential 5‐HT_2C receptor antagonists, RS102,221 (6.94) and SB242,084 (7.39), and the preferential 5‐HT_2B receptor antagonist, LY266,097 (6.66). The antipsychotic, clozapine, which had marked affinity for 5‐HT_2A receptors, blocked the recruitment of Gαq/11 by 5‐HT and DOI with pK_B values of 8.54 and 8.14, respectively. Its actions were mimicked by the “atypical” antidepressant and 5‐HT_2A receptor antagonist, mirtazapine, which likewise blocked 5‐HT and DOI‐induced Gαq/11 protein activation with pK_B values of 7.90 and 7.76, respectively. In conclusion, by use of an immunocapture/scintillation proximity strategy, this study shows that native 5‐HT_2A receptors in rat frontal cortex specifically recruit Gαq/11 and that this action is blocked by clozapine and mirtazapine. Quantification of 5‐HT_2A receptor‐mediated Gαq/11 activation in frontal cortex should prove instructive in characterizing the actions of diverse classes of psychotropic agent. Synapse 63:95–105, 2009. © 2008 Wiley‐Liss, Inc.