Nuclear and Cytoplasmic Changes in the Culex pipiens (Diptera: Culicidae) Alimentary Canal During Metamorphosis and Their Relationship to Programmed Cell Death

Abstract

The metamorphic programmed death of Culex pipiens (L.) alimentary canal cells was studied. Hoechst DNA, Acridine Orange, and TUNEL (terminal deoxynucleotidyl transferase mediated dUTP nick end labeling) staining of larval alimentary canal whole mounts showed that programmed death of polytene cells started during the 4th stadium. The cell death program was initiated in anterior stomach polytene cells and progressed to posterior stomach and gastric caeca polytene cells. Agarose gel electrophoresis confirmed that fragmentation of alimentary canal DNA into nucleosome size fragments occurred. Diploid nuclei of the anterior and posterior stomach increased in number during the time in which programmed death of polytene cells occurred. However, TUNEL staining showed that just before pupation there was also degradation of some diploid nuclear DNA in both the anterior and posterior stomach, which suggested that some of these cells also underwent programmed cell death. MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide) staining of the alimentary canal indicated that mitochondrial activity persisted while polytene nuclei were degraded. In addition, histological examination of serial thick sections showed that prepupal midgut cells had vacuoles which suggested that cytoplasmic cell death occurred during metamorphosis.