Fig. 1.

Experimental timelines. Ten days of escalating morphine injections were followed by either tissue harvesting or a withdrawal period of either a 3-day acute withdrawal or a 2-month extended withdrawal before obtaining samples. Primary neuronal cell cultures were prepared from E17 embryos on day in vitro 1 (DIV 1). On DIV 21, the media from noncontrol plates were replaced with morphine-containing media as needed. On DIV 28, naloxone, MK-801, or H-89 was added to cultures as needed. All cultures were harvested on DIV30.