Visualizing Calcium Signaling in Astrocytes

This set of movies shows calcium signaling in cultured astrocytes from the cerebral cortex of rats. These movies are intended as educational tools that may be useful in a neuroscience course or in a course focused on imaging techniques or lectures about cellular communication, calcium signaling, or the roles of glia.

Movies 1 and 2 are high-magnification and low-magnification views of astrocytes from the cerebral cortex of rats communicating spontaneously by calcium signaling in cell culture. Although astrocytes do not generate electrical impulse, as nerves do, they communicate with other glial cells and nerves by chemical signaling (1, 2). Both movies are time-lapse recordings acquired by confocal microscopy of experiments lasting approximately 20 min. Still images from each are shown as Figs. 1 and 2. The cells contain calcium indicators that allow changes in intracellular calcium concentration to be visualized as increases in brightness. Movie 1 is a high-magnification video showing that calcium is released from internal stores and also enters the cytoplasm through calcium-permeable channels in the cell membrane, and that inter- and intracellular calcium waves are evident during cell signaling in astrocytes. Movie 2 shows a low-magnification video of spontaneous signaling among astrocytes and the response to adding the neurotransmitter glutamate. In this video, glutamate activates membrane receptors that cause an increase in intracellular calcium concentration (visible as a brief flash among all cells in the field), followed by increased signaling activity among astrocytes. Waves of calcium are seen sweeping through long, slender cellular processes.

The time-lapse video in Movie 3 shows spontaneous signaling among astrocytes in coculture with dorsal root ganglion (DRG) neurons and the response of astrocytes to electrical impulses firing in DRG axons (see Fig. 3 for a still image). Axons firing electrical impulses release adenosine 5′-triphosphate (ATP) in part by a nonsynaptic and nonvesicular mechanism involving stretch-activated channels (volume-activated anion channels) (1). Calcium increases when the axons fire electrical impulses (action potentials). Action potentials cause the axons to swell, activating volume-activated anion channels that release ATP and other small molecules. The ATP stimulates purinergic receptors on astrocytes, causing an increase in intracellular calcium and stimulating the release of substances from astrocytes that stimulate calcium responses in nearby astrocytes. This signaling between axons and astrocytes can be disrupted by electrical stimulation in the presence of compounds that block the ATP-permeable volume-activated anion channels in the axon membrane. Astrocytes near synapses can also respond to neurotransmitters released by neurons at synapses (although there are no synapses in this preparation). This activates neurotransmitter receptors on astrocytes, causing a rise in intracellular calcium concentration. The time-lapse movie compresses observations of approximately 15 min.

Learning Resource Type: Video

Context: Undergraduate upper division, graduate, professional (degree program)

Intended Users: Teacher, learner

Intended Educational Use: Learn, research, teach

Discipline: Neuroscience

Keywords: neuron-glia interactions, calcium wave, glial cell, imaging, movie