Reduced Diameter, Perimeter and Cross Sectional Area of Corpus Callosum Axons Are Markers of a Deficit of Long-Range Connectivity in Autism

Background:

Autism is associated with lack of coherence, deficits in complex information processing, and intense and narrowly focused interests with a tendency to concentrate on systems that operate repetitively such as computers, games, or machines. MRI and neuropathological studies suggest that these functional anomalies are caused by defects in connectivity with short-range overconnectivity and long-range underconnectivity. Our studies of the corpus callosum in idiopathic autism revealed CC agenesis selectively affecting segments III-V in 3 of 11 examined autistic subjects, hypoplasia with a 49% deficit of axonal connections and a 37% reduction in the numerical density of axons in all agenesis-free autistic subjects (Wegiel et al 2017).

Objectives:

The aim of this postmortem study was to test the hypothesis that focal lack of CC axons and diffuse deficit of axons, reported as agenesis and hypoplasia, are associated with a third marker of long-range connectivity - reduced CC axon diameter. This pathology may directly contribute to reduced volume and capacity of information transfer between hemispheres, and lack of coherence and deficits in complex information processing in autistic subjects.

Methods: To detect markers of the deficit of long-range connectivity, 4 parameters were estimated using electron microscopy: axon diameter, perimeter, cross section area and myelin thickness in the brains of 10 subjects diagnosed with idiopathic autism and 10 brains of age and sex-matched control subjects. The brain hemispheres were fixed with formalin, dehydrated, embedded in celloidin and cut into equidistant serial 50-µm thick sections. Ultrastructural study was focused on axons in the CC genu. Equidistant serial celloidin sections were fixed with osmium tetroxide, dehydrated and embedded in Epon 812. The sections were photographed using a Hitachi 7500 electron microscope at 12,000 magnification. For each case twelve electron micrographs were used for measurements of axonal diameter, perimeter, crossectional area and myelin thickness. On average 420 axons were evaluated per case.

Results: The ultrastructural study revealed a significant (p<0.01) reduction of axon diameter by 15%, axon area by 19%, and axon perimeter by 9%. The distribution curve revealed a prevalence of small axons (less than 0.5 µm diameter), a similar percentage of middle-size axons (0.5-0.7 µm) but a reduced percentage of thick axons (>7 µm) in autistic subjects. Measurements of myelin thickness were less consistent due to the variable contribution of clusters of axons with a small diameter, very little amount axoplasm but an unusually thick myelin sheath found in some autistic subjects. Despite of these interindividual differences, the myelin thickness was reduced by 7% on average.

Conclusions: The study reveals another feature of the developmental impairment of long distance connectivity with a significant reduction in the diameter, perimeter, and cross sectional area of axons in the genu of the corpus callosum of subjects diagnosed with idiopathic autism. This pattern of pathology is considered a sign of reduced capacity and reduced volume of information transferred by long distance interhemispheric connections. The results suggest that CC sporadic agenesis, common hypoplasia and axonal developmental defects are an integral component of brain pathology contributing to the clinical autism phenotype.