Elsevier

Neuropharmacology

Volume 60, Issues 7–8, June 2011, Pages 1347-1354
Neuropharmacology

Increased serotonin axons (immunoreactive to 5-HT transporter) in postmortem brains from young autism donors

https://doi.org/10.1016/j.neuropharm.2011.02.002Get rights and content

Abstract

Imaging studies of serotonin transporter binding or tryptophan retention in autistic patients suggest that the brain serotonin system is decreased. However, treatment with drugs which increase serotonin (5-HT) levels, specific serotonin reuptake inhibitors (SSRIs), commonly produce a worsening of the symptoms. In this study we examined 5-HT axons that were immunoreactive to a serotonin transporter (5-HTT) antibody in a number of postmortem brains from autistic patients and controls with no known diagnosis who ranged in age from 2 to 29 years. Fine, highly branched, and thick straight fibers were found in forebrain pathways (e.g. medial forebrain bundle, stria terminalis and ansa lenticularis). Many immunoreactive varicose fine fibers were seen in target areas (e.g. globus pallidus, amygdala and temporal cortex). Morphometric analysis of the stained axons at all ages studied indicated that the number of serotonin axons was increased in both pathways and terminal regions in cortex from autism donors. Our findings provide morphological evidence to warrant caution when using serotonin enhancing drugs (e.g. SSRIs and receptor agonist) to treat autistic children.

This article is part of a Special Issue entitled ‘Trends in Neuropharmacology: In Memory of Erminio Costa’.

Highlights

► Increased axons reactive to 5-HT transporter in postmortem brain from autism donors. ► Increased axons in autism donors from 2.5 to 29 yr compared to age-matched controls. ► Increased axons in MFB and ansa lenticularis pathways from brainstem to forebrain. ► Increased axons seen in fusiform cortex, superior temporal cortex and amygdala. ► Caution about using drugs that increase 5-HT agonist or uptake blockers in autism.

Introduction

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders defined by social and communication deficits as well as restricted behaviors. The Centers for Disease Control states that the average prevalence of ASDs identified among children aged 8 years increased 57% in 10 sites in USA from 2002 to 2006. Incidence of ASD worldwide is not uniform, with regional variations from 1.1/1000 in China (Zhang and Ji, 2005) to 9.1/1000 in the most recent findings in the US. This is an incidence greater than both childhood cancers and diabetes combined, yet little is known about autism’s cause, prevention or treatment. Originally described as one of the most heritable mental illnesses (Bailey et al., 1995), recent re-evaluation of this description has shown methodological biases, misinterpretations, and erroneous assumptions in much of the original data (Chamak, 2010). The more likely explanation is that autism occurs in response to an environmental factor.

Elevated prenatal cortisol due to stress is associated with several negative conditions including aborted fetuses, excessive fetal activity, delayed fetal growth and development, premature delivery, low birth-weight, attention and temperament problems in infancy, externalizing problems in childhood, psychopathology, and chronic illness in adulthood (Field and Diego, 2008) and many studies indicate that increased stress (either psychological or physical) during pregnancy presents a significant risk factor for the occurrence of autism. These stressful events; such as infection, bereavement, depression, socioeconomic stress, vaginal bleeding, and threatened miscarriage may seem diverse, but all present one common endophenotype – activation of the homeostatic mechanisms of the hypothalamic–pituitary–adrenal (HPA) axis, elevating blood cortisol levels. In turn, the increase in cortisol may act as an epigenetic factor – changing the developmental trajectory of neuronal maturation necessary for normal function.

A variety of obstetrical complications have been associated with an increased risk of autism, including bleeding during pregnancy (Brimacombe et al., 2007, Juul-Dam et al., 2001), low Apgar scores (Larsson et al., 2005, Glasson et al., 2004), long labors or precipitous labor (Glasson et al., 2004), threatened abortion (Glasson et al., 2004), induced labor (Brimacombe et al., 2007, Glasson et al., 2004), and breech presentation. Psychologically, pregnant women who report sadness (Zhang et al., 2010), anxiety or adverse life events (Beversdorf et al., 2005), physical stress (including exposure to storms) (Kinney et al., 2008), were all at greater risk of giving birth to a child with autism. Stress in pregnancy causes dysregulation of fetal HPA reactivity, which endures into postnatal life, even adulthood (Egliston et al., 2007). A further indication that the HPA axis has been perturbed in autism during development, is in the number of studies which report that children with autism have altered HPA Reactivity, including lower levels of cortisol (Curin et al., 2003), altered rhythms (Corbett et al., 2008), and responses to stress (Corbett et al., 2008).

The relationship between serotonin and stress is long standing. One study conducted in the laboratory of Dr. Erminio Costa showed that stress related hormones from the adrenal gland were able to increase serotonin turnover in the brain (Azmitia et al., 1970). Using a method involving injections of radioactive tryptophan, coupled with HPLC measures of plasma and brain tryptophan and serotonin, adrenalectomy (removal of endogenous glucocorticoids) resulted in a 50% decrease in turnover that was completely reversed by injections of corticosterone. This finding has been replicated in a variety of forms and species over the last 40 years (Azmitia et al., 1993, Meijer and de Kloet, 1998, Leonard, 2005). Stress acting during early development in rodents can reduce the postsynaptic receptors of the serotonin system and glucocorticoid receptors throughout life (Veenema, 2009, Mitchell et al., 1990). Bioinformatic analysis identified nine binding sites in various serotonin receptors (HTR1D, HTR1F, HTR2A, HTR3A, and HTR6) for transcription factors in the glucocorticoid receptor family indicating a new avenue to explain stress induced serotonin activation (Falkenberg and Rajeevan, 2010). Thus, it would be expected that the reported increase in prenatal stress in autism would lead to a compromised function of the serotonin system.

A number of imaging studies indicate that the brain serotonin system in autistic patients is reduced (Chugani et al., 1997, Chugani et al., 1999, Makkonen et al., 2008). This would support the use of serotonin enhancing drugs such as SSRIs to treat this disorder (Kolevzon et al., 2006, Fatemi et al., 1998, Mehlinger et al., 1990). However, large clinical studies show that SSRIs can make autistic children worse, not better (Brodkin et al., 1997, King et al., 2009). A large meta-analysis reached a similar conclusion; there is no evidence of effect of SSRIs in children and emerging evidence of harm (Williams et al., 2010). In order to help resolve this puzzle, and facilitate the development of an effective therapy, immunocytochemical studies of serotonin neurons were performed in pathological specimens. Our results indicate a substantial increase in brain serotonin axons in young autistic donors and suggest that treatment with serotonin enhancing drugs is not indicated.

Section snippets

Brain banks

The postmortem brains were obtained from the Brain Bank for Disabilities and Aging in Staten Island, the Autism Tissue Program in Princeton New Jersey, and the NICHD Brain and Tissue Bank for Developmental Disorders at University of Maryland, Baltimore. Results of the autopsies and the Autism Diagnostic Interview-revised (ADIr) summaries are shown in Table 1 for the autism and “no known psychiatric diagnosis” (NKD) control donors. The average age for the autism group (n = 10; 14.33 yrs, range

5-HTT immunoreactive axons in fiber tracts

In humans there are two ascending pathways to the forebrain cortical structures from the serotonergic neurons in the midbrain raphe nuclei. Both pathways were heavily labeled with 5-HTT-IR axons (Fig. 1). One is the medial forebrain pathway (MFB), which receives fibers mainly from the median raphe nucleus, and the other is the ansa lenticularis (Ansa Lent), which receives fibers from the dorsal raphe nucleus through the zona incerta. The MFB projects most of its fibers to the cortex through the

Discussion

The increase of serotonin axons in the brains at all ages from autistic donors compared to NKD control donors was dramatic not only in the captured images, but also in the morphometric analysis. The increased presence of serotonin axons was seen in three of the major serotonin pathways: MFB, Ansa Lent, and Stria Terminalis. This consistent increase indicates that the results observed are not confined to one portion of the serotonin system, but rather is a global increase in all ascending

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