Ependymin, a Brain Extracellular Glycoprotein, and CNS Plasticity^a

Abstract

Ependymin, a glycoprotein of the brain ECF, has been implicated in the neurochemistry of memory and neuronal regeneration. Three behavioral experiments (swimming with a float, avoidance conditioning, and classical conditioning) in the goldfish and one in the mouse (T-maze learning) indicate that ependymin has a role in the synaptic changes that take place in the consolidation step of memory formation and the activity-dependent phase of sharpening of goldfish retinotectal connections during neuronal regeneration. The ECF concentration of the protein was found to decrease after the goldfish learned to associate a light stimulus (CS) with the subsequent arrival of a shock (US): paired CS-US gave changes whereas an unpaired presentation of CS-US gave no changes relative to the unstimulated controls. Ependymin is present in ECF as a mixture of three disulfide-linked dimers of two acidic (alpha and beta) polypeptide chains (37 kDa and 31 kDa). Upon removal of its N-linked glycan fragment by N-glycosidase F, the beta chain yields gamma-ependymin (26 kDa). Determinations of the amino acid sequence of gamma-ependymin indicate that it is a unique protein with no long sequence homologies to any known polypeptide. There are, however, small segments (5-7 amino acids long) with homologies to fibronectin, laminin, and tubulin. Ependymin has the capacity to polymerize into FIP (after activation by phosphorylation) in response to events that deplete ECF calcium. FIP is insoluble in 2% SDS in 6 M urea, 10 mM Ca2+Ac2, 100% acetic acid, chloroform/methanol (2/1), saturated KCNS, and even 100% trifluoroacetic acid. FIP was found to be present in goldfish brain and to be formed as a labeled product in vivo. Ependymin's FIP-forming property was used to propose a molecular hypothesis for generating synaptic changes in response to local extracellular depletions of calcium at sites of "associating inputs." The model assumes that, following NMDA receptor stimulation, the translocated PKC that is generated activates extracellular ependymin by converting it to its phosphorylated form using presynaptically released ATP. The hypothesis was tested in studies of LTP of rat hippocampal slices at CA1. After LTP, new sites that stained with antisera to ependymin, visible at 100x, were obtained in its potentiated radiatum in the CA1 region but not in the unpotentiated CA3. Electron microscopic studies showed that the horseradish peroxidase reaction product obtained was localized at synaptic clefts and postsynaptic regions. The results suggest that FIP may be formed at extracellular and postsynaptic loci where multiple associating inputs interact at CA1.

Plasticity of boldness in rainbow trout, Oncorhynchus mykiss: do hunger and predation influence risk-taking behaviour?

“…Risk can, however, reduce foraging without affecting appetite (Metcalfe, 1987), and since exposure to a predator induces a stress response it seems likely that upregulation of CRF and GABA A receptor in high threat conditions may represent simultaneous activation of both physiological stress response and appetitive pathways. Ependymin plays an important role in behavioural responses to the environment (Shashoua, 1991). One such role is that of memory formation (Shashoua, 1991), and variation in its expression may therefore be linked with the anticipation of events, such as predator attacks as observed in this study.…”

“…Ependymin plays an important role in behavioural responses to the environment (Shashoua, 1991). One such role is that of memory formation (Shashoua, 1991), and variation in its expression may therefore be linked with the anticipation of events, such as predator attacks as observed in this study. This effect was strongest in shy fish, which are known for reduced competitive ability compared to bold animals, an effect also linked with upregulation of ependymin (Aubin-Horth et al, 2005;Sneddon et al, 2011).…”

Extracellular matrix and its role in conveying glial/neural interactions in health and disease

“…The same Victor Shashoua referred to above, who demonstrated the excitability of a polyelectrolyte synthetic membrane, later discovered an extracellular glycoprotein, ependymin, associated with learning in fish, mice and rats (Shashoua [58]; Shashoua and Schmidt [60]; Shashoua et al [59]). He proposed that the ability of this protein to facilitate heteropolymer combination in the matrix, and its modulation by calcium, contributed to neural plasticity.…”

Over-Expression, Secondary Structure Characterization, and Preliminary X-ray Crystallographic Analysis of Xenopus tropicalis Ependymin

“…Absorptivity coefficient of ε = 1.8 (mg/mL) −1 cm −1 at λ = 280 nm was calculated by using the numbers of tyrosine and tryptophan in the protein [17]. The homogeneity of the final frog EPN was checked using SDS-PAGE under both reducing (with 5 mM DTT) and non-reducing (without DTT) conditions (Figure 2) and the aliquoted protein flash-frozen in liquid nitrogen for storage in −80 • C. 4 …”

“…Additionally, EPN being one of the abundant glycoproteins in the brain extracellular fluid and cerebrospinal fluid in the teleost fish, has been suggested to have various roles in memory consolidation, neuronal regeneration, brain calcium homeostasis [4,5] as well as in cold adaptation, and even in determining aggressiveness [6,7]. Interestingly, orthologues of fish EPN, also exist in other animals such as sea urchin, frog, and even in mammals, and have been also named as EPDR (ependymin-related) protein or MERP (mammalian ependymin-related protein).…”