Info

fyn

+

(C)

Biscardi et al., 1993

Rek

+

(R)

Fiordalisi and Maness, 1999

yes

+

(C)

Biscardi et al., 1993

R, results with retina; C, data from retinal cell cultures.

R, results with retina; C, data from retinal cell cultures.

with little serum, a large number of rods were found. Moreover, it appeared that when serum was present in the medium, rod development was somehow arrested. Further studies showed that the serum effect was indirect. It was found that serum acted by stimulating Müller cells in the culture to proliferate and release the cytokine, leukemia inhibitory factor (LIF), which in turn arrested rod development (Fig. 2.14). Similar experiments in rat retinal cultures indicate that ciliary neurotrophic factor (CNTF) and LIF inhibit rod development by redirecting rod-precursors to become bipolar cells (Kirsch et al., 1996; Ezzedine et al., 1997). Although these findings point to the exciting possibility that Müller cells regulate rod differentiation, it remains to be shown that CNTF/LIF released by Müller cells has an inhibitory effect in vivo.

There are other situations where Müller cellderived extrinsic factors appear to promote neuronal survival. Müller cell-conditioned medium has

Figure 2.14. The inhibitory effect of Müller cells on rod differentiation. (A) The histograms show the number of rhodopsin+ cells (rods) is decreased in retinal cultures that contain fetal calf serum (FCS) or Müller cells. (B) There is a dramatic decrease in the number of rods in Müller cell-conditioned medium (MCM) but not in 3T3 conditioned medium (3T3CM). (C) the suppression of rod generation in FCS can be mimicked by adding transforming growth factor-a (TGFa) which is a known Müller cell mitogen. It appears that inhibition of rod generation by FCS is an indirect effect, and arises from proliferation of Müller cells in FCS containing culture medium. Also, the conditioned medium effect could be blocked by antibodies to leukemia inhibitory factor (data not shown) which suggests that LIF released by Müller cells inhibits rod generation (Neophytou et al., 1997). (Copyright 1997 Company of Biologists Ltd., reprinted with permission.)

Figure 2.14. The inhibitory effect of Müller cells on rod differentiation. (A) The histograms show the number of rhodopsin+ cells (rods) is decreased in retinal cultures that contain fetal calf serum (FCS) or Müller cells. (B) There is a dramatic decrease in the number of rods in Müller cell-conditioned medium (MCM) but not in 3T3 conditioned medium (3T3CM). (C) the suppression of rod generation in FCS can be mimicked by adding transforming growth factor-a (TGFa) which is a known Müller cell mitogen. It appears that inhibition of rod generation by FCS is an indirect effect, and arises from proliferation of Müller cells in FCS containing culture medium. Also, the conditioned medium effect could be blocked by antibodies to leukemia inhibitory factor (data not shown) which suggests that LIF released by Müller cells inhibits rod generation (Neophytou et al., 1997). (Copyright 1997 Company of Biologists Ltd., reprinted with permission.)

been reported to support the survival of ganglion cells (Sarthy et al., 1985; Armson et al., 1987), and recent studies show that a combination of growth factors and neuronal stimulation can mimic the effects of the Müller cell-conditioned medium on ganglion cell survival (Meyer-Frank et al., 1995). This exciting observation provides good evidence that Müller cells may produce growth factors that are important for long-term neuronal survival in the retina.

2.2.6. Retinoic Acid

Retinoic acid (RA) has long been recognized as a morphogenetic factor in the developing retina (McCaffrey et al., 1991; Hyatt et al., 1996; Hyatt and Dowling, 1997). Treatment of zebrafish with RA stimulates precocious rod differentiation, and conversely, inhibition of RA synthesis in the eye leads to retarded rod development (Hyatt et al., 1996). In dissociated rat retinal cultures, RA application results in an increase in the number of progenitors that develop as rods (Stenkamp et al., 1993; Kelly et al., 1994). In agreement with these observations, the retinas of double null mice lacking the retinoic acid receptors ß2 and y2 are thinner, and show limited photoreceptor differentiation (Grondona et al., 1996).

The cellular sources of retinoic acid in the developing retina have yet to be identified. Recent studies suggest that Müller cells may be a major source

Figure 2.15. Müller cells can synthesize and secrete retinoic acid. Müller cell cultures from rabbit retina were initially incubated with [11,12-3H] all-irans-reti-nol, and the retinoid content of the media and the cells were subsequently analyzed by HPLC. The figure shows time course of accumulation of retinoic acid in medium (o), and retinaldehyde(o), retinyl esters (■J.and retinoic acid ( •) in cells. The data show that significant amounts of retinoic acid are secreted into the medium (Edwards et al., 1992). (Copyright 1992 Academic Press, Inc., reprinted with permission.)

of this retinoid. Immunocytochemical experiments, for example, show that an aldehyde dehydrogenase (ADH-2) which catalyzes the conversion of retinaldehyde to retinoic acid is present in Müller cells (McCaffrey et al., 1991). Direct evidence that Müller cells can synthesize and secrete retinoic acid was demonstrated in a recent biosynthetic study (Edwards et al., 1992). In that study, Müller cell cultures obtained from adult rabbit retina were incubated with radioactive retinol, and the cells and incubation medium were analyzed by high performance liquid chromatography (HPLC). The results showed that Müller cells synthesized both retinoic acid and retinaldehyde. Although the retinaldehyde was retained within the Müller cells, most of the retinoic acid was rapidly released into the medium (Fig. 2.15).

If retinoic acid is synthesized and secreted by Müller cells in the postnatal retina, Müller cells could exert a significant influence on the development of the late-generated retinal neurons, e.g. rods and bipolars. The observation that in vitro rod differentiation is influenced by retinoic acid (Kelley et al., 1994; Stenkamp et al., 1993) is consistent with this idea.

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