Abstract. Astroglial cells in vivo and in vitro respond to hormones, growth factors, and neurotransmitters by changing from an epithelial-like to stellate morphology. We have studied the temporal relationship between receptor activation, second messenger mobilization, and morphological changes using LRM55 astroglial cells. Maintenance of an altered morphology required continuous beta-adrenergic receptor activation. These changes appeared to be mediated by cAMP since they were elicited by its analogue, dihutyryl cAMP, and by forskolin, a direct activator of adenylate cyclase. Changes in cell morphology may require a relatively small increase in intracellular cAMP, since receptor-stimulated changes in cAMP levels were transient and peaked •5 min after receptor activation while changes in morphology took at least 30 min to reach a new steady state. Time-lapse videomicroscopy and high voltage electron microscopy indicated that receptor activation resulted in a sequence of morphological events. Time-lapse observations revealed the development and enlargement of openings through the cytoplasm associated with cytoplasmic withdrawal to the perinuclear region and process formation. Higher resolution high voltage electron microscopy indicated that the transition to a stellate morphology was preceded by the appearance of two distinct cytoplasmic domains. One contained an open network of filaments and organelles. The other was characterized by short broad cytoplasmic filaments. The first domain was similar to cytoplasm in control cells while the second was associated with the development and enlargement of openings through the cytoplasm and regions of obvious cytoplasmic withdrawal.D FFERENTIATION of astrocytes in vivo (9,24,36,37) and in vitro (8,13,14,16,21,25,32) is associated with dramatic changes in cell morphology. Cells change from an epithelial-like to a stellate morphology. Similar changes are observed in certain populations of astrocytes in the adult brain. Astrocytes in the supraoptic nucleus undergo dramatic morphological changes at parturition and after water deprivation by extending and withdrawing processes (24, 36, 37). When astrocytes in primary cell culture (3, 13-15, 21, 25, 31, 32) or C-6 glioma cells (22) are treated with beta-adrenergic agonists, activators of adenylate cyclase, or analogues of cAMP, cells change from an epitheliallike to a stellate morphology. These studies have demonstrated the occurrence of morphological changes, but have not thoroughly examined the relationships between beta-adrenergic receptor activation, intracellular CAMP, and morphology.Since activation of beta-adrenergic receptors stimulates the synthesis of cAMP in primary cell cultures of astrocytes (10,12,31,38) and in C-6 cells (6, 23), it has been proposed that changes in morphology result from activation of cAMPdependent protein kinase (7,20). Consistent with this hypothesis is the observation that activation of beta-adrenergic receptors results in the phosphorylation of the intermediate filament proteins, glial fibrilla...

We have developed a method for permeabilizing axons and reactivating the fast transport of microscopically visible organelles. Saltatory movements of organelles in motor axons isolated from lobster walking legs were observed using Nomarski optics and time-lapse video microscopy. In the center of the axon most of the particles and mitochondria moved in the retrograde direction, but immediately below the axolemma the majority moved in the anterograde direction. When axons were permeabilized with 0.02% saponin in an adenosine 5'-triphosphate (ATP)-free "internal" medium, all organelle movement ceased. Saltatory movements resembling those in intact axons immediately reappeared upon the addition of MgATP. Very slight movement could be detected with ATP concentrations as low as 10 microM, and movement appeared to be maximal with 1 to 5 mM ATP. Vanadate, which does not affect axonal transport in intact axons, inhibited the reactivated organelle movements in permeabilized axons. Movement was rapidly and reversibly inhibited by 50 to 100 microM sodium orthovanadate. The effects of vanadate, including the time course of inhibition, its reversibility, and its concentration dependence, are consistent with the hypothesis that a dyneinlike like molecule may play a role in the mechanism of fast axonal transport.