Hope it will help you
<script language='javascript'>
<!--
function EvImageOverChange(name, direction)
{
switch(direction)
{
case 'in':
name.src = "image/OverImage.gif";
break;
case 'out':
name.src = "image/InitialImage.gif";
break;
}
}
</script>
in the code behind insert this in the Page load method:
if(!IsPostBack)
{
myImageButton.ImageUrl = "image/InitialImage.gif";
myImageButton.Attributes["OnMouseOver"] =
"javascript:EvImageOverChange(this, 'in');";
myImageButton.Attributes["OnMouseOut"] =
"javascript:EvImageOverChange(this, 'out');";
}