Morphometric analysis of astrocytes in brainstem respiratory regions

Abstract

Astrocytes, the most abundant and structurally complex glial cells of the central nervous system, are proposed to play an important role in modulating the activities of neuronal networks, including respiratory rhythm-generating circuits of the preBötzinger complex (preBötC) located in the ventrolateral medulla of the brainstem. However, structural properties of astrocytes residing within different brainstem regions are unknown. In this study astrocytes in the preBötC, an intermediate reticular formation (IRF) region with respiratory-related function, and a region of the nucleus tractus solitarius (NTS) in adult rats were reconstructed and their morphological features were compared. Detailed morphological analysis revealed that preBötC astrocytes are structurally more complex than those residing within the functionally distinct neighboring IRF region, or the NTS, located at the dorsal aspect of the medulla oblongata. Structural analyses of the brainstem microvasculature indicated no significant regional differences in vascular properties. We hypothesize that high morphological complexity of preBötC astrocytes reflects their functional role in providing structural/metabolic support and modulation of the key neuronal circuits essential for breathing, as well as constraints imposed by arrangements of associated neurons and/or other local structural features of the brainstem parenchyma.

The role of PHOX2B‐derived astrocytes in chemosensory control of breathing and sleep homeostasis

“…Although brainstem astrocytes show distinct morphologies to cortical astrocytes (Sheikhbahaei et al . ), thus making the definition of brainstem gliosis challenging, we did not find an increase in astrocyte number or morphologies in the PHOX2B‐astrocyte‐ablated mice in the RTN region. We also tested the extent to which PHOX2B‐astrocyte ablation resulted in changes in the number of PHOX2B‐derived cells in the locus coeruleus, dorsal motor nucleus of the vagus nerve, and VII nucleus, all of which are known to be PHOX2B ‐derived (Fig.…”

“…; Ogata & Kosaka, ), astrocytes show highly overlapping domains (Sheikhbahaei et al . ). These highly overlapping RTN astrocyte domains preclude objective and accurate quantification by cytoplasmic glial markers.…”

“…These data align well with recent findings implicating astrocytes in the modulation of breathing circuits (Sheikhbahaei et al . ). The observation that PHOX2B ‐derived progenitor cells give rise to both astrocytes and neurons, albeit from different embryonic domains, indicates that physiologically relevant neuron‐glial units show developmental relationships in the brainstem.…”

“…Upon evaluation of the RTN region, we did not observe an appreciable decrease in GFAP-positive or ALDH1L1-positive cells. Objective quantification of astrocytes is fraught with caveats (Ostergaard & Jensen, 2013), especially in the RTN region where, unlike in the cerebrum (Bushong et al 2002;Ogata & Kosaka, 2002), astrocytes show highly overlapping domains (Sheikhbahaei et al 2018). These highly overlapping RTN astrocyte domains preclude objective and accurate quantification by cytoplasmic glial markers.…”

High‐Resolution Three‐Dimensional Imaging of Individual Astrocytes Using Confocal Microscopy

“…The same caveat applies for GFAP, the most widely used structural astrocyte marker, and for the newest small‐molecule astrocyte markers, like the cationic‐pyridinium family of markers (Preston et al., 2018; Preston, Cervasio, & Laughlin, 2019). The majority of structural astrocyte analyses are performed using GFAP exclusively (SheikhBahaei et al., 2018; Viola et al., 2009) or injectable intracellular fluorescent dyes (Butt & Ransom, 1989; Moye, Diaz‐Castro, Gangwani, & Khakh, 2019). GFAP is found in primary, secondary, and some tertiary astrocyte branches, but not in higher‐order processes, excluding significant portions of the astrocyte from analysis.…”