The cerebral cortex presents with alterations in the number of specific cell types in autism spectrum disorder (ASD). Astrocytes have many functions in the brain including a role in higher cognitive functions and in inflammatory brain processes. Therefore, an alteration in number, function, and/or activation state of astrocytes, could be present in ASD. We quantified astrocyte number in the gray and white matter of the prefrontal cortex-BA9, BA46, and BA47-in 15 ASD and 15 age- and sex-matched control cases. We labeled astrocytes with antibodies against the protein GFAP and S100β, markers of astrocytes. We found a significant decrease in the number of astrocytes in the gray and white matter of all prefrontal areas of interest with both markers. We also found an increased state of activation of GFAP+ astrocytes in all areas. A reduced number of astrocytes in the cerebral cortex in ASD could lead to impaired synaptic function and disrupted connectivity. An increased astrocyte activation may indicate a chronic mild inflammatory state of the cerebral cortex in ASD. Overall, we found that astrocytes are disrupted in ASD.

Chandelier (Ch) cells are cortical interneurons with axon terminal structures known as cartridges that synapse on the axon initial segment of excitatory pyramidal neurons. Previous studies indicate that the number of Ch cells is decreased in autism, and that GABA receptors are decreased in the Ch cell synaptic target in the prefrontal cortex. To further identify Ch cell alterations, we examined whether the length of cartridges, and the number, density, and size of Ch cell synaptic boutons, differed in the prefrontal cortex of cases with autism versus control cases. We collected samples of postmortem human prefrontal cortex (Brodmann Area (BA) 9, 46, and 47) from 20 cases with autism and 20 age- and sex-matched control cases. Ch cells were labeled using an antibody against parvalbumin, a marker that labeles soma, cartridges, and synaptic boutons. We found no significant difference in the average length of cartridges, or in the total number or density of boutons in control subjects vs. subjects with autism. However, we found a significant decrease in the size of Ch cell boutons in those with autism. The reduced size of Ch cell boutons may result in reduced inhibitory signal transmission and impact the balance of excitation to inhibition in the prefrontal cortex in autism.

Lay abstract

The cerebral cortex affected with autism spectrum disorder presents changes in the number of neurons and glia cells, possibly leading to a dysregulation of brain circuits and affecting behavior. However, little is known about cell number alteration in specific layers of the cortex in autism spectrum disorder. We found an increase in the number of neurons and a decrease in the number of astrocytes in specific layers of the prefrontal cortex in postmortem human brains from autism spectrum disorder cases. We hypothesize that this may be due to a failure in neural stem cells to shift differentiation from neurons to glial cells during prenatal brain development. These data provide key anatomical findings that contribute to the bases of autism spectrum disorder pathogenesis.

Interlaminar astrocytes (ILAs) are a subset of cortical astrocytes that reside in layer I, express GFAP, have a soma contacting the pia, and contain long interlaminar processes that extend through several cortical layers. We studied the prenatal and postnatal development of ILAs in three species of primates (rhesus macaque, chimpanzee, and human). We found that ILAs are generated prenatally likely from radial glial (RG) cells, that ILAs proliferate locally during gestation, and that ILAs extend interlaminar processes during postnatal stages of development. We showed that the density and morphological complexity of ILAs increase with age, and that ILAs express multiple markers that are expressed by RG cells (Pax6, Sox2, and Nestin), specific to inner and outer RG cells (Cryab and Hopx), and astrocyte markers (S100β, Aqp4, and GLAST) in prenatal stages and in adult. Finally, we demonstrated that rudimentary ILAs in mouse also express the RG markers Pax6, Sox2, and Nestin, but do not express S100β, Cryab, or Hopx, and that the density and morphological complexity of ILAs differ between primate species and mouse. Together these findings contribute new information on astrogenesis of this unique class of cells and suggest a lineal relationship between RG cells and ILAs.

An alteration in the balance of excitation-inhibition has been proposed as a common characteristic of the cerebral cortex in autism, which may be due to an alteration in the number and/or function of the excitatory and/or inhibitory cells that form the cortical circuitry. We previously found a decreased number of the parvalbumin (PV)+ interneuron known as Chandelier (Ch) cell in the prefrontal cortex in autism. This decrease could result from a decreased number of Ch cells, but also from decreased PV protein expression by Ch cells. To further determine if Ch cell number is altered in autism, we quantified the number of Ch cells following a different approach and different patient cohort than in our previous studies. We quantified the number of Ch cell cartridges-rather than Ch cell somata-that expressed GAT1-rather than PV. Specifically, we quantified GAT1+ cartridges in prefrontal areas BA9, BA46, and BA47 of 11 cases with autism and 11 control cases. We found that the density of GAT1+ cartridges was decreased in autism in all areas and layers. Whether this alteration is cause or effect remains unclear but could result from alterations that take place during cortical prenatal and/or postnatal development.

Interlaminar astrocytes (ILA) in the cerebral cortex possess a soma in layer I and extend an interlaminar process that runs perpendicular to the pia into deeper cortical layers. We examined cerebral cortex from 46 species that encompassed most orders of therian mammalians, including 22 primate species. We described two distinct cell types with interlaminar processes that have been referred to as ILA, that we termed pial ILA and supial ILA. ILA subtypes differ in somatic morphology, position in layer I, and presence across species. We further described rudimentary ILA that have short GFAP⁺ processes that do not exit layer I, and "typical" ILA with longer GFAP⁺ processes that exit layer I. Pial ILA were present in all mammalian species analyzed, with typical ILA observed in Primates, Scandentia, Chiroptera, Carnivora, Artiodactyla, Hyracoidea, and Proboscidea. Subpial ILA were absent in Marsupialia, and typical subpial ILA were only found in Primate. We focused on the properties of pial ILA by investigating the molecular properties of pial ILA and confirming their astrocytic nature. We found that while the density of pial ILA somata only varied slightly, the complexity of ILA processes varied greatly across species. Primates, specifically bonobo, chimpanzee, orangutan, and human, exhibited pial ILA with the highest complexity. We showed that interlaminar processes contact neurons, pia, and capillaries, suggesting a potential role for ILA in the blood-brain barrier and facilitating communication among cortical neurons, astrocytes, capillaries, meninges, and cerebrospinal fluid.