Differentiation of Recoverin Immunoreactive Cone Bipolar Cells and Their Timing of Synaptic Formation with GABAergic Amacrine Cells in the Rat Retina / 대한해부학회지

Recoverin is a member of the large family of EF-hand calcium binding proteins (Baimbridge et al., 1992), and it is thought to be involved in the regulation of phosphodiesterase in photoreceptors and in the phosphorylation of activated rhodopsin (Polans et al., 1996). Although the functional significance of recoverin in cone bipolar cells is not fully understood, the antiserum against recoverin has been widely used to identify a certain population of cone bipolar cells (Milam et al., 1993; Sasso's Pognetto et al., 1994; Euler & W sle, 1995). GABA is well known to act as major neurotransmitters in the mammalian central nervous system including retina. This study was conducted to identify the development process of recoverin-labeled cone bipolar cells, and the timing points of synaptic formation of the labeled bipolar cells and GABAergic amacrine cells in the rat retina. The results were as follows; In the adult rat retina, recoverin-labeled cone bipolar cells were subdivided into twotypes; type 2 cells with axon terminal stratified in sublamina a of the inner plexiform layer (IPL), and type 8 cells with axon terminals stratified in sublamina b of the IPL. Recoverin-labeled cone bipolar cells began to appear from postnatal day 5. The axon terminals of recoverin-labeled type 2 cone bipolar cells stratified at postnatal day 10, while those of type 8 cone bipolar cells stratified at postnatal day 13. The axon terminals of type 2 cone bipolar cells made ribbon synapses onto GABAergic amacrine cells in the IPL at postnatal day 10. These results demonstrate that recoverin-labeled type 2 cone bipolar cells differentiate earlier than recoverin-labeled type 8 cone bipolar cells, and suggest that GABAergic amacrine cells may play important roles in visual processing of recoverin-labeled type 2 cone bipolar cells by making synapse onto these cells at early stage. Synapses between type 2 cone bipolar cells and GABAergic amacrine cells are formed about the time of postnatal day 10 for visual processing.

THE EFFECT AND MECHANISM OF ASTROCYTE ON SYNAPTIC FORMATION IN RATS / 解剖学报

Objective To study the effect of astrocyte on synapse formation and the molecular mechanism. Methods Cortical astrocytes were isolated and purified from neonatal rats.On the 2h,7th day,14th day and 21th day after passage,we counted the number of astrocytes and the culture medium(astrocyte-conditioned medium,ACM)was harvested to measure the concentration of estrogen(E 2)by using ELISA techniques.Based on the model of pure culures of neonatal cortical neurons,the experimental groups were designed as follows:1.pure neuron cultures(group N);2.ACM cultures(group A);3.mixed cultures(group M);4.E 2 cultures(group E 2);5.ACM+Tamoxifen(estrogen receptor antagonist)cultures(group A+T);6.Tamoxifen cultures(group T).Then synaptic puncta in every group was stained and counted through immunofluorescence,and we also compared the differences in puncta number among those six groups(at 9th day in culture,number/per neuron). Results The numbers of astrocytes were:1?10 4/ml, 1.1?10 6/ml, 1.4?10 6/ml, 1.5?10 6/ml; The concentrations of E 2 were:(ng/L):0, 117?22, 266?22,252?27 respectively.No estrogen was detected in the primary culture medium.The concenteration of estrogen increased in correspondence with the culturing days and reached the peak around at the 14th day, then decreased gradually but kept at a certain high level,and the numbers of synaptic puncta of per neuron in group N,A,M,E 2,A+T,T were:14?3;79?5;83?8;80?6;32?3;29?3 respectively.The treatment of pure neuron culture with ACM increased the number of synapses on per neuron by up to 6 fold by comparison with pure neuronal culture.Exogenic estradiol added into pure neurons can mimick the effect of the ACM.Tamoxifen which is antagonist of estrogen receptor could decrease the effect of ACM by 75%.Conclusion The astrocytes of neonatal rat cortex do secrete E 2.Astrocyte-derived estrogen may be the molecule regulating the synaptic formation through estrogen receptors.