Degeneration of neurons, synapses, and neuropil and glial activation in a murine Atm knockout model of ataxia-telangiectasia

Abstract

Neural degeneration is one of the clinical manifestations of ataxia-telangiectasia, a disorder caused by mutations in the Atm protein kinase gene. However, neural degeneration was not detected with general purpose light microscopic methods in previous studies using several different lines of mice with disrupted Atm genes. Here, we show electron microscopic evidence of degeneration of several different types of neurons in the cerebellar cortex of 2-month-old Atm knockout mice, which is accompanied by glial activation, deterioration of neuropil structure, and both pre- and postsynaptic degeneration. These findings are similar to those in patients with ataxia-telangiectasia, indicating that Atm knockout mice are a useful model to elucidate the mechanisms underlying neurodegeneration in this condition and to develop and test strategies to palliate and prevent the disease.

Figure 1

(A) Electron micrographs of a normal Purkinje neuron in a control animal compared with a degenerating Purkinje cell in a knockout specimen (B). The degenerating neuron has both a crenated surface profile and nucleolemma, more electron dense cytoplasm, and filiform processes originating from its surface (arrows) and is surrounded by confluent vacuolated spaces (asterisks) and distended cell processes (large dots). These features are in contrast with the normal Purkinje neuron, which has a smooth perikaryal contour and nucleolemma, except for the single indentation of the latter, lighter cytoplasm, and a lesser density of mitochondria, and its processes, such as one dendrite (D), are much thicker. The control neuropil is also tight and full, lacking any significant interstitial spaces and distended processes surrounding the Purkinje neuron. PCN, Purkinje cell nuclei; GCN, granule cell nucleus. The arrow in A points to the Golgi apparatus. The magnification is the same in both panels.

Figure 3

Electron micrographs demonstrating microglial activation, neuropil disruption, and neuronal degeneration in an Atm knockout specimen. (A and B) represent two presumably consecutive phases of microglial activation associated with neuropil loss. Both panels contain a small elongated cell with numerous filiform appendages, a dark cytoplasm with lysosomes (arrow), crenated nucleolemma, and clumpy chromatin including a prominent nucleolus, i.e., features that have been correlated previously with microglia (cited in the text). Putative early stages are associated with vacuolations in the neuropil (stars) into which the filiform appendages of the microglial cells insinuate themselves (A). Presumably later stages are associated with much larger, lacune-like disruptions of the neuropil (B), which may result from the confluence of vacuoles such as those illustrated in A. C and D illustrate unequivocally degenerating neurons whose cytoplasm is severely disrupted, having lost most organelles, including mitochondria and the endoplasmic reticulum. Based on the morphological features of the normal cerebellum, the cell in C may be a Golgi neuron and that in D may be a granule neuron. Asterisks in C and D indicate vacuolations similar to those illustrated in Figs. 1 and 4, which may result from either the degeneration of the processes from the neuron in each panel or from the loss of nearby elements in the neuropil. Magnification in A and C as in D.

Figure 4

Electron micrographs demonstrating degenerating pre- and postsynaptic elements (DPrE and DPoE, respectively) in an Atm knockout specimen, recognized by swelling, loss of cytoplasmic texture, and membrane-like remains in profiles participating in synaptic junctions recognized by their synaptic clefts (curved arrows) and, in the case of presynaptic elements (A–C), by some remaining synaptic vesicles (hollow arrows, used in all panels to highlight clusters of synaptic vesicles in both normal and degenerating profiles). Degenerating postsynaptic elements can be recognized by their associated synaptic clefts (curved arrow in B) to which a presynaptic profile containing synaptic vesicles (hollow arrow) is apposed. In addition to the pre- and postsynaptic degenerating profiles, all panels display an irregular, patchy loss of texture and disruption of the neuropil, which is scattered both around the degenerating profiles as well as around the nearby, otherwise apparently normal neuropil. The most severely disrupted neuropil is entirely devoid of protoplasm and organelles, containing only membrane-like remains, and is highlighted with asterisks in all panels.

Figure 2

Electron micrographs of a degenerating neuron identifiable by synapses on its surface (curved arrows) from an Atm knockout specimen. The synapses on the perikaryon, the clear, granular cytoplasm, the prominent, elongated Golgi apparatus, the lobulated nucleus with relatively homogeneous chromatin, and the prominent nucleolus are among the features that this cell shares with Golgi neurons in the normal cerebellum. However, this neuron presents multiple, often confluent vacuolations within the cytoplasm, indicating that it is undergoing a degenerative process. GCN, granule cell nucleus.