High serum serotonin in sudden infant death syndrome.

Sudden infant death syndrome (SIDS), the leading cause of postneonatal infant mortality, likely comprises heterogeneous disorders with the common phenotype of sudden death without explanation upon postmortem investigation. Previously, we reported that ∼40% of SIDS deaths are associated with abnormalities in serotonin (5-hydroxytryptamine, 5-HT) in regions of the brainstem critical in homeostatic regulation. Here we tested the hypothesis that SIDS is associated with an alteration in serum 5-HT levels. Serum 5-HT, adjusted for postconceptional age, was significantly elevated (95%) in SIDS infants (n = 61) compared with autopsied controls (n = 15) [SIDS, 177.2 ± 15.1 (mean ± SE) ng/mL versus controls, 91.1 ± 30.6 ng/mL] (P = 0.014), as determined by ELISA. This increase was validated using high-performance liquid chromatography. Thirty-one percent (19/61) of SIDS cases had 5-HT levels greater than 2 SDs above the mean of the controls, thus defining a subset of SIDS cases with elevated 5-HT. There was no association between genotypes of the serotonin transporter promoter region polymorphism and serum 5-HT level. This study demonstrates that SIDS is associated with peripheral abnormalities in the 5-HT pathway. High serum 5-HT may serve as a potential forensic biomarker in autopsied infants with SIDS with serotonergic defects.

Dentate gyrus abnormalities in sudden unexplained death in infants: morphological marker of underlying brain vulnerability.

Sudden unexplained death in infants, including the sudden infant death syndrome, is likely due to heterogeneous causes that involve different intrinsic vulnerabilities and/or environmental factors. Neuropathologic research focuses upon the role of brain regions, particularly the brainstem, that regulate or modulate autonomic and respiratory control during sleep or transitions to waking. The hippocampus is a key component of the forebrain-limbic network that modulates autonomic/respiratory control via brainstem connections, but its role in sudden infant death has received little attention. We tested the hypothesis that a well-established marker of hippocampal pathology in temporal lobe epilepsy-focal granule cell bilamination in the dentate, a variant of granule cell dispersion-is associated with sudden unexplained death in infants. In a blinded study of hippocampal morphology in 153 infants with sudden and unexpected death autopsied in the San Diego County medical examiner's office, deaths were classified as unexplained or explained based upon autopsy and scene investigation. Focal granule cell bilamination was present in 41.2% (47/114) of the unexplained group compared to 7.7% (3/39) of the explained (control) group (p < 0.001). It was associated with a cluster of other dentate developmental abnormalities that reflect defective neuronal proliferation, migration, and/or survival. Dentate lesions in a large subset of infants with sudden unexplained death may represent a developmental vulnerability that leads to autonomic/respiratory instability or autonomic seizures, and sleep-related death when the infants are challenged with homeostatic stressors. Importantly, these lesions can be recognized in microscopic sections prepared in current forensic practice. Future research is needed to determine the relationship between hippocampal and previously reported brainstem pathology in sudden infant death.

Brainstem deficiency of the 14-3-3 regulator of serotonin synthesis: a proteomics analysis in the sudden infant death syndrome.

Impaired brainstem responses to homeostatic challenges during sleep may result in the sudden infant death syndrome (SIDS). Previously we reported a deficiency of serotonin (5-HT) and its key biosynthetic enzyme, tryptophan hydroxylase (TPH2), in SIDS infants in the medullary 5-HT system that modulates homeostatic responses during sleep. Yet, the underlying basis of the TPH2 and 5-HT deficiency is unknown. In this study, we tested the hypothesis that proteomics would uncover previously unrecognized abnormal levels of proteins related to TPH2 and 5-HT regulation in SIDS cases compared with controls, which could provide novel insight into the basis of their deficiency. We first performed a discovery proteomic analysis of the gigantocellularis of the medullary 5-HT system in the same data set with deficiencies of TPH2 and 5-HT levels. Analysis in 6 SIDS cases and 4 controls revealed a 42-75% reduction in abundance in 5 of the 6 isoforms identified of the 14-3-3 signal transduction family, which is known to influence TPH2 activity (p < 0.07). These findings were corroborated in an additional SIDS and control sample using an orthogonal MS(E)-based quantitative proteomic strategy. To confirm these proteomics results in a larger data set (38 SIDS, 11 controls), we applied Western blot analysis in the gigantocellularis and found that 4/7 14-3-3 isoforms identified were significantly reduced in SIDS cases (p ≤ 0.02), with a 43% reduction in all 14-3-3 isoforms combined (p < 0.001). Abnormalities in 5-HT and TPH2 levels and 5-HT(1A) receptor binding were associated with the 14-3-3 deficits in the same SIDS cases. These data suggest a potential molecular defect in SIDS related to TPH2 regulation, as 14-3-3 is critical in this process.

Development of brainstem 5-HT1A receptor-binding sites in serotonin-deficient mice.

The sudden infant death syndrome is associated with a reduction in brainstem serotonin 5-hydroxytryptamine (5-HT) and 5-HT(1A) receptor binding, yet it is unknown if and how these findings are linked. In this study, we used quantitative tissue autoradiography to determine if post-natal development of brainstem 5-HT(1A) receptors is altered in two mouse models where the development of 5-HT neurons is defective, the Lmx1b(f/f/p) , and the Pet-1⁻/⁻ mouse. 5-HT(1A) receptor agonist-binding sites were examined in both 5-HT-source nuclei (autoreceptors) and in sites that receive 5-HT innervation (heteroreceptors). In control mice between post-natal day (P) 3 and 10, 5-HT(1A) receptor binding increased in several brainstem sites; by P25, there were region-specific increases and decreases, refining the overall binding pattern. In the Lmx1b(f/f/p) and Pet-1⁻/⁻ mice, 5-HT(1A)-autoreceptor binding was significantly lower than in control mice at P3, and remained low at P10 and P25. In contrast, 5-HT(1A) heteroreceptor levels were comparable between control and 5-HT-deficient mice. These data define the post-natal development of 5-HT(1A)-receptor binding in the mouse brainstem. Furthermore, the data suggest that 5-HT(1A)-heteroreceptor deficits detected in sudden infant death syndrome are not a direct consequence of a 5-HT neuron dysfunction nor reduced brain 5-HT levels. To elucidate the developmental relationship between serotonin (5-HT) levels and 5-HT(1A) receptors in the brainstem, we examined 5-HT(1A) binding in two 5-HT-deficient mouse models. In nuclei containing 5-HT neurons, 5-HT(1A) binding was decreased (autoreceptors), while binding was maintained in projection sites (heteroreceptors). Thus, brainstem 5-HT(1A)-heteroreceptor-binding sites do not appear developmentally sensitive to reduced brain 5-HT levels.

Sudden and unexpected death in early life: proceedings of a symposium in honor of Dr. Henry F. Krous.

Reported here are the proceedings of a symposium given in honor of Dr. Henry F. Krous upon his retirement as Clinical Professor of Pathology and Pediatrics at the University of California Schools of Medicine, and as Director of the San Diego SIDS/SUDC Research Project. Dr. Krous' distinguished 37-year-career was dedicated to research into sudden unexpected death in infancy and childhood, notably the sudden infant death syndrome (SIDS) and sudden unexplained death in childhood (SUDC). The presentations were given at the International Conference on Stillbirth, SIDS, and infant survival on October 5, 2012, in Baltimore, MD, USA. Eight colleagues of Dr. Krous whose own professional careers were touched by his efforts discussed forensic issues related to SIDS, tissue banking, animal models in SIDS, brainstem studies in SIDS, genetic studies in SIDS, establishment of a SUDC registry, neuropathologic research in SUDC, and potential shared mechanisms underlying sudden and unexpected death in early life. The wide scope of the presentations crossed the disciplines of forensic pathology, pediatric pathology, neuropathology, neuroscience, physiology, genetics, and bereavement, and attest to Dr. Krous' far-reaching influence upon SIDS and SUDC research.

Pregnancy, delivery, and neonatal outcomes among women with incarcerated uteri: A population-based study of a large US database.

OBJECTIVE: Incarcerated uterus occurs at a rate of 1:3000 pregnancies. Previous studies focused on risk factors and management options, providing limited information about pregnancy outcomes. This study evaluates the effect of incarcerated uterus on pregnancy, delivery, and neonatal outcomes. METHODS: Retrospective study using the Healthcare Cost and Utilization Project Nationwide Inpatient Sample from 2004 to 2014. Incarcerated uterus was identified using ICD-9 code 654.3X. Multivariate logistic regression analysis was used to compare maternal and neonatal outcomes among women with and without incarcerated uterus while adjusting for confounders. RESULTS: Incarcerated uteri were identified in 370 pregnancies, and 9,096,418 pregnancies were control cases. Compared to controls, women with incarcerated uterus were more likely to be Caucasian, have smoked during pregnancy, have had a previous caesarean section, have thyroid disease, endometriosis, leiomyomas, pelvic inflammatory disease and adhesions, and ovarian cyst (P-value < 0.05 all). Women with incarcerated uterus were more likely to have placenta previa (aOR 3.1, 95% CI 1.3-7.4), deliver by caesarean section (aOR 2.4, 95% CI 1.8-3.1), have postpartum hemorrhage (aOR 2.8, 95% CI 1.8-4.4), and require blood transfusion (aOR 5.2, 95% CI 3.1-8.8). Hydronephrosis occurred more often in women with incarcerated uterus (0.8% versus 0.1%). Moreover, they were more likely to have infants with congenital anomalies (aOR 4.0, 95% CI 1.5-10.6). Rates of preeclampsia, preterm birth, and small for gestational age were similar between the two groups (P-value > 0.05, all). CONCLUSION: Women with incarcerated uterus were more likely to encounter adverse delivery and neonatal outcomes compared to the general population. These findings may help guide prenatal counseling and prenatal surveillance.

Lack of association of the serotonin transporter polymorphism with the sudden infant death syndrome in the San Diego Dataset.

Dysfunction of medullary serotonin (5-HT)-mediated respiratory and autonomic function is postulated to underlie the pathogenesis of the majority of sudden infant death syndrome (SIDS) cases. Several studies have reported an increased frequency of the LL genotype and L allele of the 5-HT transporter (5-HTT) gene promoter polymorphism (5-HTTLPR), which is associated with increased transcriptional activity and 5-HT transport in vitro, in SIDS cases compared with controls. These findings raise the possibility that this polymorphism contributes to or exacerbates existing medullary 5-HT dysfunction in SIDS. In this study, we tested the hypothesis that the frequency of LL genotype and L allele are higher in 179 SIDS cases compared with 139 controls of multiple ethnicities in the San Diego SIDS Dataset. We observed no significant association of genotype or allele with SIDS cases either in the total cohort or on stratification for ethnicity. These observations do not support previous findings that the L allele and/or LL genotype of the 5-HTTLPR are associated with SIDS.

Brainstem serotonergic deficiency in sudden infant death syndrome.

CONTEXT: Sudden infant death syndrome (SIDS) is postulated to result from abnormalities in brainstem control of autonomic function and breathing during a critical developmental period. Abnormalities of serotonin (5-hydroxytryptamine [5-HT]) receptor binding in regions of the medulla oblongata involved in this control have been reported in infants dying from SIDS. OBJECTIVE: To test the hypothesis that 5-HT receptor abnormalities in infants dying from SIDS are associated with decreased tissue levels of 5-HT, its key biosynthetic enzyme (tryptophan hydroxylase [TPH2]), or both. DESIGN, SETTING, AND PARTICIPANTS: Autopsy study conducted to analyze levels of 5-HT and its metabolite, 5-hydroxyindoleacetic acid (5-HIAA); levels of TPH2; and 5-HT(1A) receptor binding. The data set was accrued between 2004 and 2008 and consisted of 41 infants dying from SIDS (cases), 7 infants with acute death from known causes (controls), and 5 hospitalized infants with chronic hypoxia-ischemia. MAIN OUTCOME MEASURES: Serotonin and metabolite tissue levels in the raphé obscurus and paragigantocellularis lateralis (PGCL); TPH2 levels in the raphé obscurus; and 5-HT(1A) binding density in 5 medullary nuclei that contain 5-HT neurons and 5 medullary nuclei that receive 5-HT projections. RESULTS: Serotonin levels were 26% lower in SIDS cases (n = 35) compared with age-adjusted controls (n = 5) in the raphé obscurus (55.4 [95% confidence interval {CI}, 47.2-63.6] vs 75.5 [95% CI, 54.2-96.8] pmol/mg protein, P = .05) and the PGCL (31.4 [95% CI, 23.7-39.0] vs 40.0 [95% CI, 20.1-60.0] pmol/mg protein, P = .04). There was no evidence of excessive 5-HT degradation assessed by 5-HIAA levels, 5-HIAA:5-HT ratio, or both. In the raphé obscurus, TPH2 levels were 22% lower in the SIDS cases (n = 34) compared with controls (n = 5) (151.2% of standard [95% CI, 137.5%-165.0%] vs 193.9% [95% CI, 158.6%-229.2%], P = .03). 5-HT(1A) receptor binding was 29% to 55% lower in 3 medullary nuclei that receive 5-HT projections. In 4 nuclei, 3 of which contain 5-HT neurons, there was a decrease with age in 5-HT(1A) receptor binding in the SIDS cases but no change in the controls (age x diagnosis interaction). The profile of 5-HT and TPH2 abnormalities differed significantly between the SIDS and hospitalized groups (5-HT in the raphé obscurus: 55.4 [95% CI, 47.2-63.6] vs 85.6 [95% CI, 61.8-109.4] pmol/mg protein, P = .02; 5-HT in the PGCL: 31.4 [95% CI, 23.7-39.0] vs 71.1 [95% CI, 49.0-93.2] pmol/mg protein, P = .002; TPH2 in the raphé obscurus: 151.2% [95% CI, 137.5%-165.0%] vs 102.6% [95% CI, 58.7%-146.4%], P = .04). CONCLUSION: Compared with controls, SIDS was associated with lower 5-HT and TPH2 levels, consistent with a disorder of medullary 5-HT deficiency.

Serotonin-related FEV gene variant in the sudden infant death syndrome is a common polymorphism in the African-American population.

An important subset of the sudden infant death syndrome (SIDS) is associated with multiple serotonergic (5-HT) abnormalities in regions of the medulla oblongata. The mouse ortholog of the fifth Ewing variant gene (FEV) is critical for 5-HT neuronal development. A putatively rare intronic variant [IVS2-191_190insA, here referred to as c.128-(191_192)dupA] has been reported as a SIDS-associated mutation in an African-American population. We tested this association in an independent dataset: 137 autopsied cases (78 SIDS, 59 controls) and an additional 296 control DNA samples from Coriell Cell Repositories. In addition to the c.128-(191_192)dupA variant, we observed an associated single-base deletion [c.128-(301-306)delG] in a subset of the samples. Neither of the two FEV variants showed significant association with SIDS in either the African-American subgroup or the overall cohort. Although we found a significant association of c.128-(191_192)dupA with SIDS when San Diego Hispanic SIDS cases were compared with San Diego Hispanic controls plus Mexican controls (p = 0.04), this became nonsignificant after multiple testing correction. Among Coriell controls, 33 of 99 (33%) African-American and 0 of 197 (0%) of the remaining controls carry the polymorphism (c.128-(191_192)dupA). The polymorphism seems to be a common, likely nonpathogenic, variant in the African-American population.

Normative distribution of substance P and its tachykinin neurokinin-1 receptor in the medullary serotonergic network of the human infant during postnatal development.

Substance P (SP) and its tachykinin NK1 receptor (NK1R) function within key medullary nuclei to regulate cardiorespiratory and autonomic control. We examined the normative distribution of SP and NK1R in the serotonergic (5-Hydroxytryptamine, [5-HT]) network of the human infant medulla during postnatal development, to provide a baseline to facilitate future analysis of the SP/NK1R system and its interaction with 5-HT within pediatric brainstem disorders in early life. [125I] labelled Bolton Hunter SP (BH-SP) tissue receptor autoradiography (n = 15), single label immunohistochemistry (IHC) and double label immunofluorescence (IF) (n = 10) were used to characterize the normative distribution profile of SP and NK1R in the 5-HT network of the human infant medulla during postnatal development. Tissue receptor autoradiography revealed extensive distribution of SP and NK1R in nuclei intimately related to cardiorespiratory function and autonomic control, with significant co-distribution and co-localization with 5-HT in the medullary network in the normal human infant during development. A trend for NK1R binding to decrease with age was observed with significantly higher binding in premature and male infants. We provide further evidence to suggest a significant role for SP/NK1R in the early postnatal period in the modulation of medullary cardiorespiratory and autonomic control in conjunction with medullary 5-HT mediated pathways and provide a baseline for future analysis of the potential consequences of abnormalities in these brainstem neurotransmitter networks during development.

The development of the medullary serotonergic system in early human life.

The serotonergic (5-HT) neurons of the medulla oblongata are postulated to comprise a system that modulates homeostatic function in response to metabolic imbalances in the internal milieu in a state-dependent manner. In this study, we define the baseline development of the topography of the human medullary 5-HT system in 30 cases ranging from the embryonic period through infancy. We used immunocytochemical techniques with the PH8 antibody which recognizes the key 5-HT synthetic enzyme, tryptophan hydroxylase, and computer-based methods of cell quantitation. In the infant medulla, 5-HT neurons were distributed in raphé, extra-raphé, and ventral positions that place these neurons adjacent to, or intermingled with, the neurons in the lower cranial nerve nuclei and reticular formation that directly mediate respiration, upper airway reflexes, and autonomic function. Along the ventral and ventrolateral surface, 5-HT neurons formed two lateral and one midline "columns" in the rostrocaudal axis that are homologous in position to chemosensitive 5-HT neurons in rats, and that correspond in part to the classic respiratory chemosensitive fields. Serotonergic neurons comprised a subpopulation of the arcuate nucleus along the ventral surface; their short processes directly abutted the surface, suggesting a role for them in monitoring carbon dioxide levels in the cerebrospinal fluid. The medullary 5-HT system began to form in the embryo, with the raphé primordia appearing as early as 7 weeks (the earliest time-point available). By 10-12 weeks, the lateral tegmental 5-HT neurons clustered into the early primordia of extra-raphé subnuclei. By 20 weeks, the "adult-like" topography of the medullary 5-HT system was in place, with subtle (quantitative) changes occurring thereafter. Thus, protracted changes occur from the prenatal period through infancy. These data provide a foundation for 5-HT neuronal analysis in pediatric brainstem disorders, as proposed in the sudden infant death syndrome.

Medullary Serotonin Neuron Abnormalities in an Australian Cohort of Sudden Infant Death Syndrome.

Serotonin (5-hydroxytryptamine [5-HT]) neurons in the medulla oblongata project extensively to key autonomic and respiratory nuclei in the brainstem and spinal cord regulating critical homeostatic functions. Multiple abnormalities in markers of 5-HT function in the medulla in sudden infant death syndrome (SIDS) have been reported, informing the hypothesis that at least a subset of SIDS cases is caused by deficits in 5-HT function resulting in impaired homeostatic responses to potentially life-threatening events during sleep. To investigate medullary 5-HT defects in SIDS further, we undertook qualitative analysis immunohistochemical assessment of 5-HT neuron expression within the medulla of SIDS infants (n41) and nonSIDS controls (n = 28) in an independent cohort from Forensic Science South Australia. Compared with controls SIDS cases had significantly higher 5-HT neuron numbers and density in addition to significantly altered 5-HT neuron morphology. Thus, for the first time, we replicated and corroborated previous observations of a significant abnormality in medullary 5-HT neuron expression in SIDS in a separate independent SIDS cohort. This study further supports the hypothesis that medullary 5-HT defects contribute to the pathogenesis of a subset of SIDS victims and provides additional evidence of a more complex abnormality in 5-HT neuron dysfunction specifically within the different caudal and rostral medullary 5-HT domains.

Abnormalities in substance P neurokinin-1 receptor binding in key brainstem nuclei in sudden infant death syndrome related to prematurity and sex.

Sudden infant death syndrome (SIDS) involves failure of arousal to potentially life threatening events, including hypoxia, during sleep. While neuronal dysfunction and abnormalities in neurotransmitter systems within the medulla oblongata have been implicated, the specific pathways associated with autonomic and cardiorespiratory failure are unknown. The neuropeptide substance P (SP) and its tachykinin neurokinin-1 receptor (NK1R) have been shown to play an integral role in the modulation of homeostatic function in the medulla, including regulation of respiratory rhythm generation, integration of cardiovascular control, and modulation of the baroreceptor reflex and mediation of the chemoreceptor reflex in response to hypoxia. Abnormalities in SP neurotransmission may therefore result in autonomic dysfunction during sleep and contribute to SIDS deaths. [125I] Bolton Hunter SP autoradiography was used to map the distribution and density of the SP, NK1R to 13 specific nuclei intimately related to cardiorespiratory function and autonomic control in the human infant medulla of 55 SIDS and 21 control (non-SIDS) infants. Compared to controls, SIDS cases exhibited a differential, abnormal developmental profile of the SP/NK1R system in the medulla. Furthermore the study revealed significantly decreased NK1R binding within key medullary nuclei in SIDS cases, principally in the nucleus tractus solitarii (NTS) and all three subdivisions of the inferior portion of the olivo-cerebellar complex; the principal inferior olivary complex (PIO), medial accessory olive (MAO) and dorsal accessory olive (DAO). Altered NK1R binding was significantly influenced by prematurity and male sex, which may explain the increased risk of SIDS in premature and male infants. Abnormal NK1R binding in these medullary nuclei may contribute to the defective interaction of critical medullary mechanisms with cerebellar sites, resulting in an inability of a SIDS infant to illicit appropriate respiratory and motor responses to life threatening challenges during sleep. These observations support the concept that abnormalities in a multi-neurotransmitter network within key nuclei of the medullary homeostatic system may underlie the pathogenesis of a subset of SIDS cases.

Serotonergic and glutamatergic neurons at the ventral medullary surface of the human infant: Observations relevant to central chemosensitivity in early human life.

Central chemoreception is the mechanism by which the brain detects the level of carbon dioxide (CO(2)) in the arterial blood and alters breathing accordingly in order to maintain it within physiological levels. The ventral surface of the medulla oblongata (VMS) of animals has long been recognized as a site of chemosensitivity, culminating in the recent identification of chemosensitive serotonergic (5-HT) and glutamatergic (Glut) neurons in this region. In this study, we analyzed the distribution of 5-HT and Glut neurons and their receptors in the arcuate nucleus (Arc) at the VMS of the human infant, using single-and double-label immunohistochemistry with specific antibodies. We also examined the expression of astrocytes, as experimental evidence suggests that astrocytes mediate, at least in part, central chemosensitivity via 5-HT and/or Glut receptors. We identified a small number of 5-HT neurons (approximately 5% of Arc neurons), distributed over the entire extent of the VMS, a large number of Glut neurons (approximately 95% of Arc neurons) that localized almost exclusively to the medial Arc, and a large number of astrocytes distributed across the entire extent of the VMS. The Arc also contained 5-HT(1A), kainate (GluR5), and 5-HT(2A) receptors, which localized predominantly to 5-HT neurons, glutamate neurons and astrocytes, respectively. Astrocytes also expressed the vesicular glutamate transporter 2 and low levels of 5-HT(1A) and kainate (GluR5) receptors, indicating that astrocytes may store and release glutamate, possibly in response to stimulation by 5-HT and/or Glut. These observations suggest that important functional interactions exist between 5-HT, glutamate, and astrocytes in the Arc. They also support the idea that the Arc is homologous to chemosensitive zones at the VMS in experimental animals. These data are important towards delineating the role of the human Arc in modulation of homeostasis, and its dysfunction in brainstem-associated pathologies such as the sudden infant death syndrome.

Multiple serotonergic brainstem abnormalities in sudden infant death syndrome.

CONTEXT: The serotonergic (5-hydroxytryptamine [5-HT]) neurons in the medulla oblongata project extensively to autonomic and respiratory nuclei in the brainstem and spinal cord and help regulate homeostatic function. Previously, abnormalities in 5-HT receptor binding in the medullae of infants dying from sudden infant death syndrome (SIDS) were identified, suggesting that medullary 5-HT dysfunction may be responsible for a subset of SIDS cases. OBJECTIVE: To investigate cellular defects associated with altered 5-HT receptor binding in the 5-HT pathways of the medulla in SIDS cases. DESIGN, SETTING, AND PARTICIPANTS: Frozen medullae from infants dying from SIDS (cases) or from causes other than SIDS (controls) were obtained from the San Diego Medical Examiner's office between 1997 and 2005. Markers of 5-HT function were compared between SIDS cases and controls, adjusted for postconceptional age and postmortem interval. The number of samples available for each analysis ranged from 16 to 31 for SIDS cases and 6 to 10 for controls. An exploratory analysis of the correlation between markers and 6 recognized risk factors for SIDS was performed. MAIN OUTCOME MEASURES: 5-HT neuron count and density, 5-HT(1A) receptor binding density, and 5-HT transporter (5-HTT) binding density in the medullary 5-HT system; correlation between these markers and 6 recognized risk factors for SIDS. RESULTS: Compared with controls, SIDS cases had a significantly higher 5-HT neuron count (mean [SD], 148.04 [51.96] vs 72.56 [52.36] cells, respectively; P<.001) and 5-HT neuron density (P<.001), as well as a significantly lower density of 5-HT(1A) receptor binding sites (P

Serotonin Receptors in the Medulla Oblongata of the Human Fetus and Infant: The Analytic Approach of the International Safe Passage Study.

The Safe Passage Study is an international, prospective study of approximately 12 000 pregnancies to determine the effects of prenatal alcohol exposure (PAE) upon stillbirth and the sudden infant death syndrome (SIDS). A key objective of the study is to elucidate adverse effects of PAE upon binding to serotonin (5-HT) 1A receptors in brainstem homeostatic networks postulated to be abnormal in unexplained stillbirth and/or SIDS. We undertook a feasibility assessment of 5-HT1A receptor binding using autoradiography in the medulla oblongata (6 nuclei in 27 cases). 5-HT1A binding was compared to a reference dataset from the San Diego medical examiner's system. There was no adverse effect of postmortem interval ≤100 h. The distribution and quantitated values of 5-HT1A binding in Safe Passage Study cases were essentially identical to those in the reference dataset, and virtually identical between stillbirths and live born fetal cases in grossly non-macerated tissues. The pattern of binding was present at mid-gestation with dramatic changes in binding levels in the medullary 5-HT nuclei over the second half of gestation; there was a plateau at lower levels in the neonatal period and into infancy. This study demonstrates feasibility of 5-HT1A binding analysis in the medulla in the Safe Passage Study.

Serotonin transporter abnormality in the dorsal motor nucleus of the vagus in Rett syndrome: potential implications for clinical autonomic dysfunction.

Autonomic dysfunction is prevalent in girls with Rett syndrome, an X-chromosome-linked disorder of mental retardation resulting from mutations in the gene encoding methyl-CpG-binding protein 2 (MeCP2). This gene plays a role in regulating neuronal activity-dependent gene expression, including brain-derived neurotrophic factor (BDNF), which is a potent serotonergic (5-HT) neuronal growth factor. We analyzed selected parameters of the 5-HT system of the medulla in autopsied patients with Rett syndrome because of the role of BDNF in 5-HT cell development and because 5-HT plays a key role in modulating autonomic control. 5-HT neurons were identified by immunostaining for tryptophan hydroxylase, the biosynthetic enzyme for 5-HT. We quantitated the number of 5-HT cells in the medulla at 2 standardized levels in 11 Rett and 7 control cases. There was no significant difference in 5-HT cell number between the groups. We analyzed binding to the serotonin transporter (SERT) using the radioligand [(125)I]-RTI-55 with tissue autoradiography in 7 Rett and 5 controls in 9 cardiorespiratory-related nuclei. In the dorsal motor nucleus of the vagus (DMX) (preganglionic parasympathetic outflow), SERT binding for the control cases decreased significantly over time (p = 0.049) but did not change in the Rett cases (p = 0.513). Adjusting for age, binding between the Rett and control cases differed significantly in this nucleus (p = 0.022). There was a marginally significant age versus diagnosis interaction (p = 0.06). Thus, altered 5-HT innervation and/or uptake in the DMX may contribute to abnormal 5-HT modulation of this major autonomic nucleus in patients with Rett syndrome. These data suggest hypotheses concerning 5-HT modulation of vagal function for testing in MeCP2 knockout mice to understand mechanisms underlying autonomic dysfunction in patients with Rett syndrome.

Differential development of 5-HT receptor and the serotonin transporter binding in the human infant medulla.

Tissue receptor autoradiography with 3H-lysergic acid diethylamide (3H-LSD), 3H-8-hydroxy-2-[di-N-propylamine] tetralin (3H-8-OH-DPAT), and 125I-RTI-55 was used to map the distribution and developmental profile of 5-HT(1A-1D) and 5-HT2 receptors, 5-HT1A receptors, and the serotonin (5-HT) transporter (SERT), respectively, to nuclei with cardiorespiratory function in the human medulla from midgestation to maturity. The distribution pattern of the 5-HT markers was heterogeneous, with variable densities of binding of each observed both in nuclei with and without 5-HT cell bodies. The highest density of binding for each marker was observed in the raphé nuclei, the site of the highest density of 5-HT cell bodies. A significant reduction in 5-HT receptor binding measured with 3H-LSD was observed between midgestation and infancy, and between infancy and maturity in multiple nuclei, but no changes were observed across infancy. A significant increase in 5-HT1A receptor binding density was observed across infancy in the hypoglossal nucleus (regression slope coefficient = 0.008 +/- 0.002, P = 0.02), and a marginally significant increase was observed in the raphé obscurus (regression slope coefficient = 0.061 +/- 0.026 [mean +/- SEM], P = 0.05). No significant age-related changes in SERT binding were observed at any time. With the exception of the hypoglossal nucleus, where 5-HT1A receptor binding increases while SERT binding remains stable, the medullary 5-HT markers analyzed in the study are essentially "in place" at birth. This study provides important baseline data that serve as a foundation for future work in pediatric 5-HT brainstem disorders, including sudden infant death syndrome.

Serotonin metabolites in the cerebrospinal fluid in sudden infant death syndrome.

Forensic biomarkers are needed in sudden infant death syndrome (SIDS) to help identify this group among other sudden unexpected deaths in infancy. Previously, we reported multiple serotonergic (5-HT) abnormalities in nuclei of the medulla oblongata that help mediate protective responses to homeostatic stressors. As a first step toward their assessment as forensic biomarkers of medullary pathology, here we test the hypothesis that 5-HT-related measures are abnormal in the cerebrospinal fluid (CSF) of SIDS infants compared with those of autopsy controls. Levels of CSF 5-hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA), the degradative products of 5-HT and dopamine, respectively, were measured by high-performance liquid chromatography in 52 SIDS and 29 non-SIDS autopsy cases. Tryptophan (Trp) and tyrosine (Tyr), the substrates of 5-HT and dopamine, respectively, were also measured. There were no significant differences in 5-HIAA, Trp, HVA, or Tyr levels between the SIDS and non-SIDS groups. These data preclude the use of 5-HIAA, HVA, Trp, or Tyr measurements as CSF autopsy biomarkers of 5-HT medullary pathology in infants who have died suddenly and unexpectedly. They do, however, provide important information about monoaminergic measurements in human CSF at autopsy and their developmental profile in infancy that is applicable to multiple pediatric disorders beyond SIDS.

Serotonin gene variants are unlikely to play a significant role in the pathogenesis of the sudden infant death syndrome.

Sudden infant death syndrome (SIDS) is defined as the sudden and unexpected death of an infant less than 12 months of age that is related to a sleep period and remains unexplained after a complete autopsy, death scene investigation, and review of the clinical history. The cause of SIDS is unknown, but a major subset of SIDS is proposed to result from abnormalities in serotonin (5-HT) and related neurotransmitters in regions of the lower brainstem that result in failure of protective homeostatic responses to life-threatening challenges during sleep. Multiple studies have implicated gene variants that affect different elements of 5-HT neurotransmission in the pathogenesis of these abnormalities in SIDS. In this review I discuss the data from these studies together with some new data correlating genotype with brainstem 5-HT neurochemistry in the same SIDS cases and conclude that these gene variants are unlikely to play a major role in the pathogenesis of the medullary 5-HT abnormalities observed in SIDS.