Astrocyte syncytium: from neonatal genesis to aging degeneration

Modern neuroscience began from all reaching and fierce conflict between “neuronismo and reticulismo” – between neuronal and reticular theories of the organization of the nervous system; the conflict culminated in December of 1906 in Stockholm where Santiago Ramon y Cajal (the proponent of the neuronal doctrine) and Camillo Golgi (who advocated the syncytial reticular organization of neural networks) delivered their Noble prize lectures (Verkhratsky, 2009). The neuronal doctrine eventually was victorious and dominated 20^th-century neuroscience and neurology. As frequently happens in science, the views of both Cajal and Golgi were correct, and as we know now the central nervous system (CNS) comprises highly coordinated networks of synaptically connected neurons and gap junction-connected neuroglia, the latter being the syncytial or reticular portion of the nervous tissue (Kiyoshi and Zhou, 2019).

Extensive gap junctional coupling is a distinctive feature of astroglia, of which astrocytes (protoplasmic, fibrous, velate, marginal, etc., for classification, see Verkhratsky and Butt, 2023) are the most numerous. Gap junctional coupling assembles astrocytes into syncytia critical for brain function (Ma et al., 2016; Kiyoshi et al., 2018). Astrocyte syncytia uniformly distribute in the CNS with certain regional differences; astrocytes can also couple with oligodendroglia and ependymoglia into panglial syncytia (Kiyoshi et al., 2018; Verkhratsky and Nedergaard, 2018). Nonetheless, exactly how this glial network contributes to the information processing in the CNS remains unknown. In part, the lingering of this question is due to the lack of knowledge and experimental approaches to monitor the functional state of an astrocyte syncytium and its effects on neuronal performance. This long-standing question has been addressed by a recent developmental study where the transition of uncoupled newborn astrocytes to a coupled syncytial network in the mouse hippocampus was scrutinized and minutely characterized (Zhong et al., 2023). Developmental evolution of a functional syncytium progresses through a coordinated maturation of astrocyte cellular morphology, the spatial organization of astrocytes and their territorial domains, gap junctional coupling, and expression of background K⁺ permeability. Further, the functional state of a developing syncytium was quantitatively assessed by its capacity for intra-syncytial equilibration of K⁺ ions that reach a state of initial maturity at P15. Furthermore, this study has provided a conceptual framework as well as suggested a suitable experimental approach for future examination of astrocyte function at the syncytial level, which represents a higher tier of organizational hierarchy.

Recent studies in both rodent and human brains suggest that the morpho-functional integration achieved by astrocytic syncytia is reversed in senescent CNS, thus contributing to age-induced cognitive decline and age-dependent neuropathologies (Popov et al., 2021, 2022; Verkhratsky et al., 2021). This emerging field of research highlights the importance of in-depth analysis of the loss-of-function of the astrocyte syncytia in the aged and diseased brain. In this Perspective article, we will review these studies and discuss their implication for future research in this area.

Astrocyte syncytium – a functional reticular system in the CNS: Astrocytes are connected through intercellular channels, connexons (Cx43, Cx30, and Cx26 representing individual subunits or connexins, which may assemble as homo or heteromers, with CX43 being the most abundant in astroglia) into an astrocytic network. Functionally, system-wide wiring of astrocyte syncytium is essential for the movements and distribution of ions, metabolites, and signaling molecules across the brain (Verkhratsky and Nedergaard, 2018). The syncytial coupling also equalizes the membrane potentials fluctuations of individual astrocytes resulting from regional neuronal activities; such equalization is termed syncytial isopotentiality, which is an astrocytic mechanism whereby a strong and constant driving force is shared among astrocytes for high-efficiency performance of numerous homeostatic transporters and ion channels, including those responsible for buffering of extracellular K⁺ and neurotransmitters clearance in the course of neuronal activation and synaptic transmission (Ma et al., 2016; Kiyoshi and Zhou, 2019). New evidence indicates that uncoupling astrocytes and hence removing syncytial isopotentiality impair synaptic transmission and plasticity. Thus, it is necessary for astrocytes to act as a syncytium to sustain brain function.

Available tools for revealing the functional state of an astrocyte syncytium: A variety of methods are available to examine the functional state, conductance and selectivity of gap junctions as well as syncytial continuity and inter-syncytial diffusion, including electrical and dye coupling assays, as well as imaging of the propagating Na⁺, Ca²⁺, and glucose waves. A powerful electrophysiological method combined with a computational model developed to examine the integrity of astrocyte syncytium allows precise characterization of its functional state. Specifically, Na⁺ (or Cs⁺) electrode solution-based electrophysiological recordings can be used for measuring the strength of astrocyte syncytial isopotentiality, as well as the capacity of a syncytium for equalization of K⁺ ions in various brain regions, including protoplasmic astrocytes in grey matters, fibrous astrocytes in the white matter, and specialized Bergmann glia and velate astrocytes in the cerebellum (Ma et al., 2016; Kiyoshi et al., 2018; Zhong et al., 2023). In the postnatal developing hippocampus, these methods were successfully employed as accurate readouts of the gain-of-function of astrocyte syncytium progressively emerging in the early postnatal developing hippocampus (Figure 1). Thus, these experimental tools should be equally powerful to interrogate the age-dependent alteration of astrocyte syncytium. Potentially, a loss-of-function of astrocyte syncytium over the course of brain aging would be an appealing hypothesis to be tested in the future.

Figure 1.

Genesis and degeneration of an astrocyte syncytium over the course of brain development and aging.

During the neonatal development (Newborn → Adult), a functional astrocyte syncytium is established at P15 in the mouse hippocampus; a process that is involved in the (1) maturation in the cellular structure and syncytial organization of astrocytes; (2) switch from expression of voltage-gated K⁺ channels (IK_a and IK_d) to predominantly open K⁺ channels (IK_open), and (3) transition of astrocytes from uncoupled individuals to syncytial coupling network. The coordinated maturation of these anatomical and functional features generated a syncytium with high efficiency for intra-syncytium equilibration of K⁺ ions. In the aging brain (Senile), astrocyte atrophy and weakened syncytial coupling are indicated by recent studies, which put forward a hypothesis that progressive reversion of the genesis process occurs in aging astrocyte syncytia. I_K, [K⁺]_p and I_K, [Na⁺]_p respectively refer to the K⁺ currents recorded with K⁺- and Na⁺-based electrode solutions. V_M, [Na⁺]_p refers to membrane voltage recorded with Na⁺-based electrode solution. I_K, endogenous, and I_K, syncytium respectively refer to the K⁺ currents generated from the endogenous K⁺ channels of the same astrocyte and those from syncytial coupled astrocytes. Modified from Figure 9 in Zhong et al. (2023) with permission.

Checklist of astrocyte properties for making a functional astrocyte syncytium: While defining/revealing the functional state is a natural first step to characterize syncytium, it is equally important to know the cause of the syncytium malfunction. In the developmental study by Zhong et al. (2023), several morphological and functional features were considered as the defining features for making an operational syncytium in the context equilibration of K⁺ ions. These include (1) an adequate morphology of astrocytes to make gap junctional coupling with 7–9 nearest neighboring astrocytes (Kiyoshi et al., 2018; Aten et al., 2022b); (2) the need to lower the interastrocytic resistance to ~4.2 MΩ through gap junctional coupling, a prerequisite for syncytial isopotentiality (Ma et al., 2016); and (3) an abundant expression of open K⁺ channels for generating and sustaining hyperpolarized membrane potentials to be shared throughout coupled astrocytes in an isopotential network. These features are not only identified/validated from the developmental studies (Zhong et al., 2023), atrophy of astrocyte in an animal model of depression (Aten et al., 2022a), and reduced expression of gap junctions in animals after ablation of microglia (Du et al., 2022) all resulted in an impaired syncytial function. Thus, in the aging and diseased brains, these astrocytic features should be the blueprints to explore further in pathological mechanistic studies.

Physiological aging, neurodegeneration, and neuropsychiatric disorders are associated with prominent morphological atrophy of astrocytes (Popov et al., 2021, 2022; Verkhratsky et al., 2021), which inevitably impairs the syncytial coupling and reduces isopotentiality of the astrocytic syncytium. This, in turn, impacts on all aspects of astrocytic homeostatic support with particular impairment of glutamate clearance and K⁺ buffering, which are powerful modulators of synaptic transmission and plasticity (Verkhratsky et al., 2021). Thus, aberrant synaptic transmission, observed in all these pathologies may result from syncytial uncoupling; while targeting astrocytic reticulum and preserving syncytial isopotentiality may appear as a valid therapeutic strategy.

This work was sponsored by a grant from the National Institute of Neurological Disorders and Stroke: RO1NS116059 (to MZ).