AmOA1 in the Honey Bee Brain Figure 3. AmOA1 is expressed in the antennal lobes, mushroom body and lateral protocerebrum. A–C: Example of cell bodies and neuropil labeled with AmOA1 in the antennal lobe. The lateral side of the antennal lobe is on the right. A: In the antennal nerve (AN), the tracts of olfactory receptor neurons (T1–T2) have a few beaded processes positive for staining with AmOA1 antibodies (arrow). The glomeruli surrounding the entrance of the antennal nerve show different patterns of AmOA1 immunoreactivity. The double arrows point out an example of low intensity AmOA1 staining in the glomerular cortex where the endings of the olfactory receptor neurons are located. In contrast, the core region of this glomerulus has a high level of AmOA1 staining. B: Neurons in the lateral group (LG) exhibit a high intensity of AmOA1 immunoreactivity in their cell bodies and in their axons that are in the coarse area (arrow) of the antennal lobes. The cells bodies of the dorsal group (DG) also show intense staining. C: The medial dorsal group (MDG) of antennal neurons was also stained with high intensity with anti-AmOA1 antibodies. D–H: AmOA1 immunoreactivity in the mushroom body. D: One half of a calyx stained with AmOA1 antiserum. Kenyon cell bodies (K) that supply the collar or outer basal ring exhibit differing AmOA1 immunoreactivity. The dendrites of Kenyon cells are labeled with high intensity in the outer wedge basal ring zone of the calyx (o.w.br) whereas less intense staining is observed in the calyx collar (co). Arrowheads indicate endings of extrinsic (feedback) neurons expressing AmOA1 in the calyx lip, the outer basal ring (o.br), and with a few branches in the collar neuropil. The inner basil ring (i. br) show relatively low levels of staining. E: A sagittal section made through the mushroom body vertical (V) and medial (M) lobes. Dorsal is at the top. In the medial and vertical lobes, there are two layers of Kenyon cell axons stained with anti-AmOA1 antibodies (arrowheads). These layers correspond to the basal ring and collar Kenyon cells. The class II clawed Kenyon cells, which have their axons in the ventral part of the vertical lobe (the c lobe), are also positive for AmOA1 expression. Arrows show the axons of c lobe Kenyon cells brightly labeled with anti-AmOA1. The cell bodies of these cells lay outside of calyx (see arrow in H). F: A frontal section through the vertical lobe of mushroom body. AmOA1 immunoreactive extrinsic neurons enter and branch in the c lobe. The arrow indicates an example of AmOA1 immunoreactive neurites of feedback neurons that enter in the vertical lobe. G: A schematic representation of a calycal cross section illustrating calycal regions receiving olfactory (yellow), gustatory and mechanosensory (dark blue), and visual input from the lobula (dark red) and medulla (light blue). H: The AmOA1 immunoreactive cell bodies of neurons located in a cluster between the dorso-medial edge of the medulla (Me) and the lateral calyx are neurons that connect the mushroom body calyx with the medulla. The cells in this cluster have fibers in the anterior superior optical tract (asot). The arrowhead shows axons from these neurons entering the medulla serpentine layer. AmOA1 positive fibers run along the dorsal edge of the lobula (Lo) and enter into the serpentine layer of the medulla. A few AmOA1 positive fibers enter through the lobula optical tract (lot) that connects lobula and mushroom bodies. I: The lateral protocerebrum (LP) exhibits few fibers or stained structures (midline is on the left). There are diffuse processes in the lateral horn (LH) that express AmOA1 in this animal. Scale bar: 50 mm in A,B,C; 25 mm in D,E,F,H; 75 mm in I. doi:10.1371/journal.pone.0014536.g003 II Kenyon cells, also referred to as ‘‘clawed’’ Kenyon cells, have soma that lie outside the calyces. Their neurites extend through the outer wall of the calyx where they produce distinctive ‘‘clawed’’ dendrites. Class II Kenyon cells have axons which supply the ventral part, or c division, of the vertical lobe and may also bifurcate to supply the vertical and medial lobes [57–60]. The calyces are divided into modality specific zones – lip, collar and basal ring - that receive input from different sensory modalities [54,60–63]. The lip receives input from olfactory projection neurons, the collar receives input from visual, tactile and gustatory processing neuropils, and the basal ring receives input from olfactory, gustatory, tactile and visual neuropils (see schematic in Fig. 3G). It is important to note here that Kenyon cell bodies were labeled with different intensity across the calyx (Figures 3D,H, 7B, D2), which is in accordance with in situ analysis [43]. The AmOA1 PLoS ONE | www.plosone.org 6 January 2011 | Volume 6 | Issue 1 | e14536