Digital, three-dimensional average shaped atlas of the heliothis virescens brain with integrated gustatory and olfactory neurons

Abstract

We use the moth Heliothis virescens as model organism for studying the neural network involved in chemosensory coding and learning. The constituent neurons are characterised by intracellular recordings combined with staining, resulting in a single neuron identified in each brain preparation. In order to spatially relate the neurons of different preparations a common brain framework was required. We here present an average shaped atlas of the moth brain. It is based on 11 female brain preparations, each stained with a fluorescent synaptic marker and scanned in confocal laser-scanning microscope. Brain neuropils of each preparation were manually reconstructed in the computer software Amira, followed by generating the atlas using the Iterative Shape Average Procedure. To demonstrate the application of the atlas we have registered two olfactory and two gustatory interneurons, as well as the axonal projections of gustatory receptor neurons into the atlas, visualising their spatial relationships. The olfactory interneurons, showing the typical morphology of inner-tract antennal lobe projection neurons, projected in the calyces of the mushroom body and laterally in the protocerebral lobe. The two gustatory interneurons, responding to sucrose and quinine respectively, projected in different areas of the brain. The wide projections of the quinine responding neuron included a lateral area adjacent to the projections of the olfactory interneurons. The sucrose responding neuron was confined to the suboesophageal ganglion with dendritic arborisations overlapping the axonal projections of the gustatory receptor neurons on the proboscis. By serving as a tool for the integration of neurons, the atlas offers visual access to the spatial relationship between the neurons in three dimensions, and thus facilitates the study of neuronal networks in the Heliothis virescens brain. The moth standard brain is accessible at

Antennal-lobe tracts in the noctuid moth, Heliothis virescens: new anatomical findings

“…The anti-SYNORF1 (Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA, USA) was raised against fusion proteins composed of glutathione-Stransferase and the Drosophila SYN1 protein (SYNORF1; Klagges et al 1996). The specificity of this antibody has been described by Klagges et al (1996), and it is reported to detect synaptic neuropil in various insect species, heliothine moths included (Berg et al 2002;Kvello et al 2009;Zhao et al 2014). After analyzing the mass-stained brains by confocal laser scanning microscopy, the preparation was rehydrated through a decreased ethanol series (10 min each) and rinsed in PBS.…”

Implementation of pigment‐dispersing factor‐immunoreactive neurons in a standardized atlas of the brain of the cockroach Leucophaea maderae

“…A comparative volumetric analysis indicates that L. maderae relies more heavily on chemosensory than on visual information than all other species studied (Drosophila melanogaster: Rein et al, 2002 Kvello et al, 2009), and that L. maderae has relatively smaller optic lobes, the largest antennal lobe, and the largest mushroom body. Chemosensory input appears to be the main input to the mushroom bodies, as suggested by linear regression analysis (Table 3).…”

“…Selective neuropil staining by the SYNORF1 antibody has previously been demonstrated in several insect species (Brandt et al, 2005;Kurylas et al, 2008;el Jundi et al, 2009;Kvello et al, 2009). It was raised against a 66-kDa fusion protein comprising glutathione-S-transferase (GST) fused to the first amino acids of the presynaptic vesicle protein synapsin I coded by its 5 0 fragment (nucleotide sequence 621-1967, shown in Fig.…”

“…The VIB protocol has been used to create standardized brains of the fruit fly Drosophila (Rein et al, 2002;Jenett et al, 2006), the desert locust Schistocerca gregaria (Kurylas et al, 2008), and the giant sphinx moth Manduca sexta (el Jundi et al, 2009), whereas the ISA method has been used in the honeybee (Rohlfing et al, 2001;Brandt et al, 2005), the desert locust (Kurylas et al, 2008), and the noctuid moth Heliothis virescens (Kvello et al, 2009). Whereas the VIB protocol can be used to determine time-or stimulus-dependent differences in the individual brains, the ISA protocol is designed to eliminate these differences to obtain an average brain for implementation of identified neurons.…”

Heterogeneity in dendritic morphology of moth antennal lobe projection neurons

“…Ordinary glomeruli in the moth AL have been analyzed in detail, and an identification method for the glomeruli has been established (Berg et al, 2002;Sadek et al, 2002;Masante-Roca et al, 2002;Greiner et al, 2004;Skiri et al, 2005;Kazawa et al, 2009;Couton et al, 2009;Varela et al, 2009), as in the cases of other insects (Galizia et al, 1999;Laissue et al, 1999). Determination of the morphology of PNs in moths by using glomeruli identification has been reported with relatively small data sets (up to 40; Sun et al, 1997;Vickers et al, 1998;Sadek et al, 2002;Kanzaki et al, 2003;Han et al, 2005;Rø et al, 2007;Kvello et al, 2009;Løfaldli et al, 2010;Zhao and Berg, 2010). In the present study, we analyzed the morphology of a total of 246 PNs, including PNs innervating ordinary glomeruli, with the glomerular identification method by using a digital atlas .…”