Primary Commissure Pioneer Neurons in the Brain of the Grasshopper Schistocerca gregaria: Development, Ultrastructure, and Neuropeptide Expression PETER LUDWIG, 1 LES WILLIAMS, 1 DICK R. NA ¨ SSEL, 2 HEINRICH REICHERT, 3 AND GEORGE BOYAN 1 * 1 Zoologisches Institut, Ludwig-Maximilians-Universita ¨t, 80333 Mu ¨ nchen, Germany 2 Department of Zoology, Stockholm University, S-10691 Stockholm, Sweden 3 Zoologisches Institut, CH-4051 Basel, Switzerland ABSTRACT The bilaterally paired primary commissure pioneer neurons in the median domain of the grasshopper brain are large, descending interneurons that uniquely express the TERM-1 antigen, even in the adult. After pioneering the primary interhemispheric brain commissure, these neurons extend TERM-1-immunoreactive collaterals into most parts of the brain except the mushroom bodies. In this report, the authors show that the TERM-1 antigen is located in the cell body cytoplasm of these neurons and not on the membranes. Screening with antisera to insect neuropeptides reveals that an antiserum recognizing peptides of the leucokinin family labels the cell body cytoplasm of the primary commissure neurons. Leucokinin-related peptides are known to modulate motility of visceral muscle, play a role in diuresis, and are likely to be neuromodulators in the insect nervous system. The primary commissure neurons differ ultrastructurally from median neurosecretory cells in that their cell body cytoplasm is more extensive, contains high numbers of mitochondria and extensive endoplasmic reticu- lum, but does not contain neurosecretory granules. In the adult, the cell somata are enveloped by multiple glia membranes and associated trophospongia. According to these ultrastructural characteristics, the primary commissure pioneers are not classical neurosecretory cells. J. Comp. Neurol. 430:118 –130, 2001. © 2001 Wiley-Liss, Inc. Indexing terms: axogenesis; primary commissure pioneer cells; leucokinin-1; TERM-1; insects In recent years, the combination of molecular, histolog- ical, and electrophysiological techniques has greatly in- creased our understanding of the development and func- tion of insect nervous systems. Several such studies involve identified neurons in the insect brain (Krautham- mer, 1985; Na ¨ ssel et al., 1992; Bacon et al., 1995; Stern et al., 1995; Lundquist et al., 1998), and, in some cases, it has proven possible to establish a clear function for the specific neurons involved (Agui, 1979, 1980; Copenhaver and Tru- man, 1986; Gammie and Truman, 1997; McNabb et al., 1999; Renn et al., 1999; Zitnan et al., 1999). Despite these advances, only small numbers of neurons in the insect brain can be shown to be identifiable based on the fact that they are the only neurons that express a given molecule (Truman and Copenhaver, 1989; Thomp- son et al., 1991; Meier et al., 1993). Neurons of the grass- hopper brain, such as the TERM-1-expressing primary commissure pioneer (PCP) cells (Meier et al., 1993; Boyan et al., 1995a; Ludwig et al., 1999) or the vasopressin-like- immunoreactive (VPLI) cells of the subesophageal gan- glion (Tyrer et al., 1993), belong to this category. Their specific antigen expression allows them to be easily iden- tified at all developmental stages, and, because they are Grant sponsor: Deutsche Forschungsgemeinschaft; Grant number: BO 1434/1-3; Grant sponsor: Swedish Natural Science Research Council. *Correspondence to: Prof. Dr. George Boyan, Zoologisches Institut der LMU, Luisenstrasse 14, 80333 Mu ¨ nchen, Germany. E-mail: boyan@zi.biologie.uni-muenchen.de Received 23 August 2000, Revised 18 October 2000; Accepted 18 October 2000 THE JOURNAL OF COMPARATIVE NEUROLOGY 430:118 –130 (2001) © 2001 WILEY-LISS, INC.