EGR-1 during activity-dependent plasticity of developing visual cortex

S324

CHARACTERIZATION

741

AND PARTIAL PURIFICATION OF NEUROPSIN-

BINDING PROTEIN

TOMOHIRO

KAMACHI,

KEIKO KATO, KAZUMASA

MATSUMOTO,

TAKUYA

YOSHIDA,

AND SADAO SHIOSAKA

Department

of Structural Cell Biology, Nara Institute of Science and Technology,

OKA, SHIGETAKA

8916-5 Takayama-cho,

Ikoma,

Nara 630-0101 To determine binding-proteins

a series of cascades concerning to neuropsin

with development

(NP). Homogenates

was in 0.15 M NaCl soluble fraction.

of mice hippocampus

R-NP; recombinant

NaCl soluble fraction were mixed and immunoprecipitated, formation of the complexes

of neuronal plasticity.

we focused the purification

and cerebral cortex were fractionated;

prepared from baculo virus expression then two complexes;

possible that the binding-protein partially for the binding-protein

in 260Kd had the function concerning in 260Kd; the combination

native-NP

system, and the 0. I5M

65Kd and 260Kd, were determined.

resulted in decrease of the enzyme activity of r-NP to 50 %I; it was possible

proteins were substrate and inhibitor against NP. Next, 260Kd-complex

of the

was only in the presence

The

that the binding-

of r-activated-NP;

it was

with the enzyme activity of NP. Now we purified

of ResourceQ

and Butyl Sepharose

4 Fast Flow resulted in the

purity to about lOO-fold.

742

INVOLVEMENT OF PROTEIN PHOSPHATASES AND KINASES IN THE INDUCTION OF FAST AND SLOW LONG-TERM POTENTIATIONS IN DEVELOPING RAT VISUAL CORTEX.

TOHRU KUROTANI.

Dept. of Physiology,

XUE-LONG

JIN AND KEISUKE TOYAMA

Kyoto Prefectural

Univ. of Med., Kawaramachi-Hirokoji,

Kamigyo-ku,

Kyoto

602-0841,

Japan

We demonstrated that theta burst stimulation produces NMDA receptor-dependent ‘fast’ long-term potentiation (fLTP) and NMDA receptor-independent L-type voltage gated calcium channel-dependent ‘slow’ LTP (sLTP) of field potentials in layer 4 of developing rat visual cortical slices evoked by white matter stimulation. fLTP, showing a fast time course, is independent of protein or RNA synthesis whereas sLTP, demonstrating a slow time course, depends upon the macromolecular syntheses (Kurotani et al., 1996). In the present study, we examined whether protein kinases and phosphatases are involved in the induction of these LTPs. fLTP and sLTP were studied separately in the presence of an Ltype voltage gated calcium channel antagonist, nimodipine, and an NMDA receptor antagonist, m-APV, respectively. Bath application of either a protein phosphatase 2B inhibitor, FK-506, or a protein phosphatase li2A inhibitor, Calyculin A, reduced the incidence of fLTP. In contrast, FK-506 increased the incidence of sLTP. On the other hand, a bath application of serine/threonine kinase inhibitor staurosporine reduced the incidence of sLTP in a dose-dependent manner but did not affect fLTP. Neither FK-506 or staurosporine, if applied after theta burst stimulation, showed any significant effects on either of the LTPs. These results suggest that fLTP requires the activation of protein phosphatases for induction while sLTP requires that of serineithreonine kinases instead. Supported by a Grant-in-Aid for Ecncouragement of Young Scientists (09780771).

743

THE ROLE OF ZIF268/EGR-1 DEVELOPING

NOBUKO

DURING ACTIVITY-DEPENDENT

PLASTICITY

OF

VISUAL CORTEX

MATAGA’, BRIAN G. CONDIE*, SAYAKA FUJISHIMA’ AND TAKAO K. HENSCH’

‘Lab for Neuronal Circuit Development, Brain Science Institute, RIKEN, Saitama 351-0198 JAPAN, *Inst. Molecular Medicine & Genetics, Medical College Georgia, Augusta, GA 30912 USA Experience-dependent plasticity in the mammalian visual cortex is a well-documented phenomenon, however, the molecular and cellular basis for this plasticity remains unknown. Recent studies suggest a possible involvement of the immediate early gene (IEG) zif268/Egr-1 in synaptic development and plasticity of cat visual cortex. We, therefore, examined a role for this IEG in developing visual cortex with mice carrying a targeted disruption of the zif268/Egr-1 gene. We first analyzed the developmental pattern of zif268/egr-I mRNA expression in wild type mice by Northern blot hybridization. In mouse visual cortex, as in other species such as monkey, cat and rat, zif268/Egr-I mRNA expression dramatically increased with eye opening or 30 min. photic stimulation following a 5 day period of dark rearing. These results demonstrated that expression of the zif268 IEG is regulated by visual input. Next, single unit recordings were obtained from the binocular zone contralateral to an eye deprived of vision. We initially confirmed that visual cortical responses developed normally in zif268/egr-I knockout (KO) mice [contralateral bias index (CBI)=0.75&).01, n=3]. A significant reduction in CBI was found in both wild type (CBI=0.43+0.04, n=8) and KO mice (CBI=0.42+0.04, n=9) that had been monocularly deprived for 4-15 days during the developmental critical period. These results indicate that the zif268 IEG is not essential for the regulation of experiencedependent plasticity in mouse visual cortex.