Shuangxia decoction alleviates p-chlorophenylalanine induced insomnia through the modification of serotonergic and immune system.

As a Traditional Chinese Medicine (TCM), Shuangxia Decoction (SXD) has been used to treat insomnia in oriental countries for more than thousands of years and it presents remarkable clinical effects. However, its active pharmacological fraction and the mechanism of sedative-hypnotic effects have not been explored. In this paper, we investigated active pharmacological fraction and revealed the detailed mechanisms underlying the sedative-hypnotic effects of SXD. It showed that SXD water extract compared to ethanol extract possessed better sedative effects on locomotion activity in normal mice and increased sleep duration in subhypnotic dose of sodium pentobarbital-treated mice. SXD alleviated p-chlorophenylalanine (PCPA) -induced insomnia by increasing the content of 5-HT in cortex [F (4, 55) = 12.67], decreasing the content of dopamine (DA) and norepinephrine (NE). Furthermore, SXD enhanced the expression of 5-HT1A and 5-HT2A receptors in hypothalamic and reduced serum levels of IL-1,TNF-α [F (5, 36) = 15.58]. In conclusion, these results indicated that SXD produced beneficial sedative and hypnotic bioactivities mediated by regulating the serotonergic and immune system.

Role of the Serotonin Receptor 7 in Brain Plasticity: From Development to Disease.

Our knowledge on the plastic functions of the serotonin (5-HT) receptor subtype 7 (5-HT7R) in the brain physiology and pathology have advanced considerably in recent years. A wealth of data show that 5-HT7R is a key player in the establishment and remodeling of neuronal cytoarchitecture during development and in the mature brain, and its dysfunction is linked to neuropsychiatric and neurodevelopmental diseases. The involvement of this receptor in synaptic plasticity is further demonstrated by data showing that its activation allows the rescue of long-term potentiation (LTP) and long-term depression (LTD) deficits in various animal models of neurodevelopmental diseases. In addition, it is becoming clear that the 5-HT7R is involved in inflammatory intestinal diseases, modulates the function of immune cells, and is likely to play a role in the gut-brain axis. In this review, we will mainly focus on recent findings on this receptor's role in the structural and synaptic plasticity of the mammalian brain, although we will also illustrate novel aspects highlighted in gastrointestinal (GI) tract and immune system.

Targeted inhibition of serotonin type 7 (5-HT7) receptor function modulates immune responses and reduces the severity of intestinal inflammation.

Mucosal inflammation in conditions ranging from infective acute enteritis or colitis to inflammatory bowel disease is accompanied by alteration in serotonin (5-hydroxytryptamine [5-HT]) content in the gut. Recently, we have identified an important role of 5-HT in the pathogenesis of experimental colitis. 5-HT type 7 (5-HT7) receptor is one of the most recently identified members of the 5-HT receptor family, and dendritic cells express this receptor. In this study, we investigated the effect of blocking 5-HT7 receptor signaling in experimental colitis with a view to develop an improved therapeutic strategy in intestinal inflammatory disorders. Colitis was induced with dextran sulfate sodium (DSS) or dinitrobenzene sulfonic acid (DNBS) in mice treated with selective 5-HT7 receptor antagonist SB-269970, as well as in mice lacking 5-HT7 receptor (5-HT7(-/-)) and irradiated wild-type mice reconstituted with bone marrow cells harvested from 5-HT7(-/-) mice. Inhibition of 5-HT7 receptor signaling with SB-269970 ameliorated both acute and chronic colitis induced by DSS. Treatment with SB-269970 resulted in lower clinical disease, histological damage, and proinflammatory cytokine levels compared with vehicle-treated mice post-DSS. Colitis severity was significantly lower in 5-HT7(-/-) mice and in mice reconstituted with bone marrow cells from 5-HT7(-/-) mice compared with control mice after DSS colitis. 5-HT7(-/-) mice also had significantly reduced DNBS-induced colitis. These observations provide us with novel information on the critical role of the 5-HT7 receptor in immune response and inflammation in the gut, and highlight the potential benefit of targeting this receptor to alleviate the severity of intestinal inflammatory disorders such as inflammatory bowel disease.

Serotonin skews human macrophage polarization through HTR2B and HTR7.

Besides its role as a neurotransmitter, serotonin (5-hydroxytryptamine, 5HT) regulates inflammation and tissue repair via a set of receptors (5HT(1-7)) whose pattern of expression varies among cell lineages. Considering the importance of macrophage polarization plasticity for inflammatory responses and tissue repair, we evaluated whether 5HT modulates human macrophage polarization. 5HT inhibited the LPS-induced release of proinflammatory cytokines without affecting IL-10 production, upregulated the expression of M2 polarization-associated genes (SERPINB2, THBS1, STAB1, COL23A1), and reduced the expression of M1-associated genes (INHBA, CCR2, MMP12, SERPINE1, CD1B, ALDH1A2). Whereas only 5HT(7) mediated the inhibitory action of 5HT on the release of proinflammatory cytokines, both 5HT(2B) and 5HT(7) receptors mediated the pro-M2 skewing effect of 5HT. In fact, blockade of both receptors during in vitro monocyte-to-macrophage differentiation preferentially modulated the acquisition of M2 polarization markers. 5HT(2B) was found to be preferentially expressed by anti-inflammatory M2(M-CSF) macrophages and was detected in vivo in liver Kupffer cells and in tumor-associated macrophages. Therefore, 5HT modulates macrophage polarization and contributes to the maintenance of an anti-inflammatory state via 5HT(2B) and 5HT(7), whose identification as functionally relevant markers for anti-inflammatory/homeostatic human M2 macrophages suggests their potential therapeutic value in inflammatory pathologies.

Relationship between serotonin and mast cells: inhibitory effect of anti-serotonin.

Serotonin (5-HT) is an important neurotransmitter that acts in both central and peripheral nervous system, and has an impact on cell proliferation, migration and apoptosis. 5HT exerts its effects via several receptors. Treatment with anti-5-HT receptors diminish the severity of contact allergy in experimental animals, an effect mediated by mast cells; while an agonist reduces the stress level and relieves pruritus in patients with atopic dermatitis. Mast cells are important for both innate and adaptive immunity and they are activated by cross-linking of FceRI molecules, which are involved in the binding of multivalent antigens to the attached IgE molecules, resulting in a variety of responses including the immediate release of potent inflammatory mediators. Serotonin is present in murine mucosal mast cells and some authors reported that human mast cells may also contain serotonin, especially in subjects with mastocytosis. Here we report the interrelationship between mast cells, serotonin and its receptor inhibitor.

Serotonin modulation of macrophage polarization: inflammation and beyond.

Macrophages display a ample plethora of effector functions whose acquisition is promoted by the surrounding cytokine and cellular environment. Depending on the stimulus, macrophages become specialized ("polarized") for either pathogen elimination, tissue repair and wound healing or immunosuppression. This "polarization" versatility allows macrophages to critically contribute to tissue homeostasis, as they promote initiation and resolution of inflammatory responses. As a consequence, deregulation of the tissue macrophage polarization balance is an etiological agent of chronic inflammation, autoimmune diseases, cancer and even obesity and insulin resistance. In the present review we describe current concepts on the molecular basis and the patho-physiological implications of macrophage polarization, and describe its modulation by serotonin (5-HT), a neurotransmitter that regulates inflammation and tissue repair via a large set of receptors (5-HTR1-7). 5-HT modulates the phenotypic and functional polarization of macrophages, and contributes to the maintenance of an anti-inflammatory state mainly via 5-HTR2B and 5-HTR7, whose activation has a great impact on macrophage gene expression profile. The identification of 5-HTR2B and 5-HTR7 as functionally-relevant polarization markers suggests their therapeutic value in inflammatory pathologies as well as their potential involvement in linking the immune and nervous systems.

[Serotonergic regulation of the immune system].

The paper presents evidence on the important contribution of the peripheral serotonin system in the process of immunomodulation. The main components of the system - serotonin, receptors and serotonin transporter - are described. Possible mechanisms of serotonin regulation of activity of immune cells - lymphocytes, macrophages and dendritic cells - are reviewed.

Serotonin Signalling in Flatworms: An Immunocytochemical Localisation of 5-HT7 Type of Serotonin Receptors in Opisthorchis felineus and Hymenolepis diminuta.

The study is dedicated to the investigation of serotonin (5-hydroxytryptamine, 5-HT) and 5-HT7 type serotonin receptor of localisation in larvae of two parasitic flatworms Opisthorchis felineus (Rivolta, 1884) Blanchard, 1895 and Hymenolepis diminuta Rudolphi, 1819, performed using the immunocytochemical method and confocal laser scanning microscopy (CLSM). Using whole mount preparations and specific antibodies, a microscopic analysis of the spatial distribution of 5-HT7-immunoreactivity(-IR) was revealed in worm tissue. In metacercariae of O. felineus 5-HT7-IR was observed in the main nerve cords and in the head commissure connecting the head ganglia. The presence of 5-HT7-IR was also found in several structures located on the oral sucker. 5-HT7-IR was evident in the round glandular cells scattered throughout the larva body. In cysticercoids of H. diminuta immunostaining to 5-HT7 was found in flame cells of the excretory system. Weak staining to 5-HT7 was observed along the longitudinal and transverse muscle fibres comprising the body wall and musculature of suckers, in thin longitudinal nerve cords and a connective commissure of the central nervous system. Available publications on serotonin action in flatworms and serotonin receptors identification were reviewed. Own results and the published data indicate that the muscular structures of flatworms are deeply supplied by 5-HT7-IR elements. It suggests that the 5-HT7 type receptor can mediate the serotonin action in the investigated species and is an important component of the flatworm motor control system. The study of the neurochemical basis of parasitic flatworms can play an important role in the solution of fundamental problems in early development of the nervous system and the evolution of neuronal signalling components.

Serotonin and dopamine receptors profile on peripheral immune cells from patients with temporal lobe epilepsy.

The role of inflammation and immune cells has been demonstrated in neurological diseases, including epilepsy. Leukocytes, as well as inflammatory mediators, contribute to abnormal processes that lead to a reduction in seizure threshold and synaptic reorganization. In this sense, identifying different phenotypes of circulating immune cells is essential to understanding the role of these cells in epilepsy. Immune cells can express a variety of surface markers, including neurotransmitter receptors, such as serotonin and dopamine. Alteration in these receptors expression patterns may affect the level of inflammatory mediators and the pathophysiology of epilepsy. Therefore, in the current study, we evaluated the expression of dopamine and serotonin receptors on white blood cells from patients with temporal lobe epilepsy with hippocampal sclerosis (TLE-HS). Blood samples from 17 patients with TLE-HS and 21 controls were collected. PBMC were isolated and stained ex vivo for flow cytometry. We evaluated the expression of serotonin (5-HT1A, 5-HT1B, 5-HT2, 5-HT2B, 5-HT2C, 5-HT3, 5-HT4), and dopamine receptors (D1, D2, D3, D4, and D5) on the cell surface of lymphocytes and innate immune cells (monocytes and granulocytes). Our results demonstrated that innate cells and lymphocytes from patients with TLE-HS showed high mean fluorescent intensity (MFI) for 5-HT1A, 5-HT1B, 5-HT2A, and 5-HT4 compared to controls. No difference was observed for 5-HT2B. For dopamine receptors, the expression of D1, D2, D4, and D5 receptors was higher on innate cells from patients with TLE-HS when compared to controls for the MFI. Regarding lymphocytes population, D2 expression was increased in patients with TLE-HS. In conclusion, there are alterations in the expression of serotonin and dopamine receptors on immune blood cells of patients with TLE-HS. Although the biological significance of these findings still needs to be further investigated, these changes may contribute to the understanding of TLE-HS pathophysiology.

Role of serotonin receptor signaling in cancer cells and anti-tumor immunity.

Serotonin or 5-hydroxytryptamine (5-HT) is a neurotransmitter known to affect emotion, behavior, and cognition, and its effects are mostly studied in neurological diseases. The crosstalk between the immune cells and the nervous system through serotonin and its receptors (5-HTRs) in the tumor microenvironment and the secondary lymphoid organs are known to affect cancer pathogenesis. However, the molecular mechanism of - alteration in the phenotype and function of - innate and adaptive immune cells by serotonin is not well explored. In this review, we discuss how serotonin and serotonin receptors modulate the phenotype and function of various immune cells, and how the 5-HT-5-HTR axis modulates antitumor immunity. Understanding how 5-HT and immune signaling are involved in tumor immunity could help improve therapeutic strategies to control cancer progression and metastasis.

Antibodies to Signaling Molecules and Receptors in Alzheimer's Disease are Associated with Psychomotor Slowing, Depression, and Poor Visuospatial Function.

BACKGROUND: Alzheimer's disease (AD) is associated with several antibodies as well as signaling molecules and receptors. These may be detrimental in the presence of a disrupted blood-brain barrier (BBB). OBJECTIVE: To investigate whether the levels of antibodies toward 33 signaling molecules involved in neurotransmitter, vascular, and immune functions were associated with AD and, within the AD group; cognitive function and mood. METHODS: Antibodies in sera from patients with mild AD [(n = 91) defined as a Mini-Mental State Examination ≥ 20 or a Clinical Dementia Rating Scale≤1] and healthy controls (n = 102) were measured with enzyme-linked immunosorbent assays. Levels in AD and controls were compared by Mann-Whitney test. In the AD group, associations between antibodies and psychometric test scores were analyzed by robust regression. The false discovery threshold was set to 0.05. RESULTS: Antibodies to serotonin receptors [5-HT2AR (effect size (r) = 0.21, p = 0.004), 5-HT2CR (r = 0.25, p = 0.0005) and 5-HT7R (r = 0.21, p = 0.003)], vascular endothelial growth factor receptor 1 [VEGFR1 (r = 0.29, p < 0.001)] and immune-receptors (Stabilin-1 (r = 0.23, p = 0.001) and C5aR1 (r = 0.21, p = 0.004) were higher in AD. Psychomotor speed was associated with D1R-abs (ß 0.49, p < 0.001), depression with ETAR-abs (ß 0.31, p < 0.001), and visuospatial function with 5-HT1AR-abs (ß 0.27, p = 0.004) despite similar antibody levels compared to controls. CONCLUSIONS: Antibody levels to VEGFR1, serotonergic receptors, and receptors in the immune system were increased in AD. Antibodies at similar levels as in controls were associated cognitive dysfunction and depression in AD.

Visualization of 5-HT Receptors Using Radioligand-Binding Autoradiography.

Described in this unit are techniques to visualize the majority of serotonin (5-hydroxytryptamine, 5-HT) receptor subtypes in sections of frozen brain tissue using receptor autoradiography. Protocols for brain extraction and sectioning, radioligand exposure, autoradiogram generation, and data quantification are provided, as are the optimal incubation conditions for the autoradiographic visualization of receptors using agonist and antagonist radioligands. © 2016 by John Wiley & Sons, Inc.

Serotonin and the 5-HT(2B) receptor in the development of enteric neurons.

We tested the hypothesis that 5-HT promotes the differentiation of enteric neurons by stimulating a developmentally regulated receptor expressed by crest-derived neuronal progenitors. 5-HT and the 5-HT(2) agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine(.)HCl (DOI) enhanced in vitro differentiation of enteric neurons, both in dissociated cultures of mixed cells and in cultures of crest-derived cells isolated from the gut by immunoselection with antibodies to p75(NTR). The promotion of in vitro neuronal differentiation by 5-HT and DOI was blocked by the 5-HT(1/2) antagonist methysergide, the pan-5-HT(2) antagonist ritanserin, and the 5-HT(2B/2C)-selective antagonist SB206553. The 5-HT(2A)-selective antagonist ketanserin did not completely block the developmental effects of 5-HT. 5-HT induced the nuclear translocation of mitogen-activated protein kinase. This effect was blocked by ritanserin. mRNA encoding 5-HT(2A) and 5-HT(2B) receptors was detected in the fetal bowel (stomach and small and large intestine), but that encoding the 5-HT(2C) receptor was not. mRNA encoding the 5-HT(2B) receptor and 5-HT(2B) immunoreactivity were found to be abundant in primordial [embryonic day 15 (E15)-E16] but not in mature myenteric ganglia. 5-HT(2B)-immunoreactive cells were found to be a subset of cells that expressed the neuronal marker PGP9.5. These data demonstrate for the first time that the 5-HT(2B) receptor is expressed in the small intestine as well as the stomach and that it is expressed by enteric neurons as well as by muscle. It is possible that by stimulating 5-HT(2B) receptors, 5-HT affects the fate of the large subset of enteric neurons that arises after the development of endogenous sources of 5-HT.

Tissue-specific self-peptides bound by major histocompatibility complex class I molecules of a human pancreatic beta-cell line.

The process of beta-cell destruction in IDDM is mediated, in part, by CD8+ T-cells. Structural characterization of HLA-I-bound self-peptides presented by the human beta-cell line HP-62 was performed to identify possible tissue-specific autoantigens in the context of CD8+ T-cell/HLA-I interactions. The sequences of the beta-cell line HLA-I-bound peptides were compared with sequence databases. Six of the obtained sequences showed homology to known precursor proteins, three of which--GLUT2 receptor, phosphatidylinositol-glycan-specific phospholipase D, and 5-hydroxytryptamine-1F receptor--have a limited, tissue-specific expression. These HLA-bound self-peptides may be part of a pool of autoantigens recognized by beta-cell reactive cytotoxic T-cells.

The 5-HT receptor cell is a new member of secondary mesenchyme cell descendants and forms a major blastocoelar network in sea urchin larvae.

A gene encoding the serotonin (5-hydroxytryptamine, 5-HT) receptor (5-HT-hpr) was identified from the sea urchin, Hemicentrotus pulcherrimus. Partial amino acid sequence deduced from the cDNA showed strong similarity to Aplysia californica 5-HT2 receptor. Immunoblotting analysis of this 5-HT-hpr protein (5-HT-hpr) with an antibody raised against a deduced peptide showed two bands. Their relative molecular masses were 69 and 53 kDa, respectively. The larger band alone disappeared after N-glycopeptidase F digestion, indicating the protein was N-glycosylated. Immunolocalization analysis showed that cells expressing the 5-HT-hpr (SRC) first appeared near the tip of the archenteron in 33-h post-fertilization (33 hpf) prism larvae. Their cell number doubled in 2 h, and 5-HT-hpr protein expression increased further without cell proliferation. SRC spread ventrally on the basal surface of the oral ectoderm in 36 hpf prism larvae, and then clockwise on the ventral ectoderm to the posterior region to complete formation of the SRC network in 48 hpf early plutei. The SRC network was comprised of 7 main tracts: 4 spicule system-associated tracts and 3 spicule system-independent tracts. The network extended short fibers to the larval body surface through the ectoderm, implicating a signal transmission system that receives exogenous signal. Double-stain immunohistochemistry with antibodies to primary mesenchyme cells showed that SRC were not stained by the antibody. In embryos deprived of secondary mesenchyme cell (SMC) by microsurgery, the number of SRC decreased considerably. These two data indicate that SRC are SMC descendants, adding a new member to the SMC lineage.

Immunomodulatory effects mediated by serotonin.

Serotonin (5-HT) induces concentration-dependent metabolic effects in diverse cell types, including neurons, entherochromaffin cells, adipocytes, pancreatic beta-cells, fibroblasts, smooth muscle cells, epithelial cells, and leukocytes. Three classes of genes regulating 5-HT function are constitutively expressed or induced in these cells: (a) membrane proteins that regulate the response to 5-HT, such as SERT, 5HTR-GPCR, and the 5HT3-ion channels; (b) downstream signaling transduction proteins; and (c) enzymes controlling 5-HT metabolism, such as IDO and MAO, which can generate biologically active catabolites, including melatonin, kynurenines, and kynurenamines. This review covers the clinical and experimental mechanisms involved in 5-HT-induced immunomodulation. These mechanisms are cell-specific and depend on the expression of serotonergic components in immune cells. Consequently, 5-HT can modulate several immunological events, such as chemotaxis, leukocyte activation, proliferation, cytokine secretion, anergy, and apoptosis. The effects of 5-HT on immune cells may be relevant in the clinical outcome of pathologies with an inflammatory component. Major depression, fibromyalgia, Alzheimer disease, psoriasis, arthritis, allergies, and asthma are all associated with changes in the serotonergic system associated with leukocytes. Thus, pharmacological regulation of the serotonergic system may modulate immune function and provide therapeutic alternatives for these diseases.

The role of serotonin and its receptors in activation of immune responses and inflammation.

Serotonin or 5-hydroxytryptamine (5-HT) is a neurotransmitter and hormone that contributes to the regulation of various physiological functions by its actions in the central nervous system (CNS) and in the respective organ systems. Peripheral 5-HT is predominantly produced by enterochromaffin (EC) cells of the gastrointestinal (GI) tract. These gut-resident cells produce much more 5-HT than all neuronal and other sources combined, establishing EC cells as the main source of this biogenic amine in the human body. Peripheral 5-HT is also a potent immune modulator and affects various immune cells through its receptors and via the recently identified process of serotonylation. Alterations in 5-HT signalling have been described in inflammatory conditions of the gut, such as inflammatory bowel disease. The association between 5-HT and inflammation, however, is not limited to the gut, as changes in 5-HT levels have also been reported in patients with allergic airway inflammation and rheumatoid arthritis. Based on searches for terms such as '5-HT', 'EC cell', 'immune cells' and 'inflammation' in pubmed.gov as well as by utilizing pertinent reviews, the current review aims to provide an update on the role of 5-HT in biological functions with a particular focus on immune activation and inflammation.

Monoclonal antibodies against the S2-serotonin receptor from rat brain that cross-react with dopamine and opiate receptors.

Balb/c mice were immunized with rat striatal integral membrane proteins. After hybridization of splenocytes with myeloma cells, hybridoma lines secreting antibodies against serotonin, dopamine and opiate receptors were detected by inhibition of ligand binding to brain membrane preparations. Antibodies from two positive lines, Mab/a9 and Mab/a18, were able to inhibit ligand binding to the S2-serotonin (Kd range: 10-100 nM), the mu-opiate (Kd range: 0.4-3 microM) and the delta-opiate receptors (Kd range: 0.7-1.1 microM), while Mab/a9 was also found to inhibit ligand binding to the D2/D4-dopamine receptor (Kd approximately 50 nM). An apparent molecular mass of 60 kDa could be ascribed to the delta-opiate receptor and apparent molecular masses of 29 and 36 kDa to the mu-opiate receptor by ligand elution from immuno-precipitates.

Cellular and subcellular distribution of the serotonin 5-HT2A receptor in the central nervous system of adult rat.

Light and electron microscope immunocytochemistry with a monoclonal antibody against the N-terminal domain of the human protein was used to determine the cellular and subcellular localization of serotonin 5-HT2A receptors in the central nervous system of adult rat. Following immunoperoxidase or silver-intensified immunogold labeling, neuronal, somatodendritic, and/or axonal immunoreactivity was detected in numerous brain regions, including all those in which ligand binding sites and 5-HT2A mRNA had previously been reported. The distribution of 5-HT2A-immunolabeled soma/dendrites was characterized in cerebral cortex, olfactory system, septum, hippocampal formation, basal ganglia, amygdala, diencephalon, cerebellum, brainstem, and spinal cord. Labeled axons were visible in every myelinated tract known to arise from immunoreactive cell body groups. In immunopositive soma/dendrites as well as axons, the 5-HT2A receptor appeared mainly cytoplasmic rather than membrane bound. Even though the dendritic labeling was generally stronger than the somatic, it did not extend to dendritic spines in such regions as the cerebral and piriform cortex, the neostriatum, or the molecular layer of the cerebellum. Similarly, there were no labeled axon terminals in numerous regions known to be strongly innervated by the immunoreactive somata and their axons (e.g., molecular layer of piriform cortex). It was concluded that the 5-HT2A receptor is mostly intracellular and transported in dendrites and axons, but does not reach into dendritic spines or axon terminals. Because it has previously been shown that this serotonin receptor is transported retrogradely as well as anterogradely, activates intracellular transduction pathways and intervenes in the regulation of the expression of many genes, it is suggested that one of its main functions is to participate in retrograde signaling systems activated by serotonin.

Synaptic plasticity of 5-hydroxytryptamine-immunoreactive terminals in the phrenic nucleus following spinal cord injury: a quantitative electron microscopic analysis.

The present study was conducted to examine the plasticity of 5-hydroxytryptamine (5-HT)-immunoreactive terminals in the rat phrenic nucleus following an ipsilateral C2 spinal cord hemisection and 30-day survival period. A retrograde horseradish peroxidase (HRP) labeling technique was used to identify the phrenic motoneurons at the electron microscopic (EM) level. After employing a pre-embedding immunocytochemical technique, the ultrastructural characteristics of 5-HT-immunoreactive terminals were qualitatively and then quantitatively analyzed with a computerized morphometric system before and after injury in separate groups of rats. The results indicated that the majority of the 5-HT-labeled terminals formed axodendritic contacts, but some 5-HT-labeled terminals made axosomatic contacts. 5-HT terminals were associated with either asymmetrical or symmetrical synapses, and some displayed postsynaptic dense bodies. Approximately 2% of the 5-HT terminals had dense-core vesicles. Although the total number of labeled and unlabeled terminals in the phrenic nucleus was reduced after hemisection, the number of 5-HT terminals in the hemisected group was greater than that of the control group. Moreover, the total number and length of asymmetrical and symmetrical synaptic active zones per 5-HT terminal were significantly greater after injury. Finally, the total number of 5-HT terminals with multiple synapses was significantly greater in the hemisected group as compared to controls. It is possible that 5-HT synaptic plasticity may be part of the morphological substrate for the unmasking of the latent crossed phrenic pathway which mediates recovery of the ipsilateral hemidiaphragm paralyzed by C2 spinal cord hemisection.