Referee 2's review (Hans-Joachim Pflüger):

Review Vömel and Wegener

Biogenic amines are major modulators of behavior in both invertebrate and vertebrate animals. The discovery of the presence of particular biogenic amines in mammals which were once thought to exclusively exist in invertebrates, makes them increasingly interesting for a wider community of scientists. This article describes for Drosophila larvae the distribution of key enzymes for the synthesis pathways of biogenic amines, and examines some of the dendritic projections of selected neurons with respect to longitudinal tracts and published knowledge on the structure of neuropils. In addition, the manuscript tries to identify functional compartments of selected neurons by genetic manipulations. It provides a very useful description of neurons in Drosophila larvae that contain the respective enzymes required for the synthesis of some biogenic amines. Less convincing is the attempt to describe the projections of neurites (dendrites) of particular neurons with respect to compartments of the neuropil as it seems that the knowledge of the (functional) structure of the neuropil is less advanced in Drosophila than in larger insects. In particular, the descriptions of neuron morphologies should be indicated in more detail in the figures which are largely free of further explanations.

Specific questions and remarks

Page 2, Introduction, first chapter
I find the statement incorrect that in insects biogenic amines are primarily synthesized in interneurons and released in the CNS. This may be true for neurons that release 5-HT or DA but is not correct for neurons releasing octopamine. Although, the term biogenic amines is commonly used for these kind of amines, I would prefer to treat each of them separately.

Page 2, Introduction, second chapter
There is an impressive body of work that describes the projections and distributions of octopaminergic neurons in larger insects. Therefore, in this whole paragraph more reference should be given to what is known from other insects than Drosophila.

Page 5 and 6
To understand all figures, in particular all transverse sections, a schematic drawing of one segmental ganglion in which all transversal projections (TP 1-4) are indicated, is absolutely required. In addition, all abbreviations used for tracts and other features of a ganglion should be contained in the figure legend. Please explain all colour codes in figure legends, for example, green shaded areas.

Page 6, first chapter
The authors claim tat it is possible to follow the neuritic projections of neurons. Therefore, the authors should explain where the serotonergic somata in neuromere a8 send their neurites.

Page 7 and figure 4
On what basis are the three midline neurons classified as unpaired? To me it looks like one is unpaired and there is one pair of neurons. Although I know that the schematic drawings used in Figure 4 (and in all other figures!) are popular among "Drosophilists", I do not find them particularly helpful, as they never show which neurites exactly run contralaterally, or ipsilateraly, or have bilaterally symmetrical projections and to which somata they really belong. Please indicate neurite paths uneqivocally.

In Figure C4 no primary neurite is drawn at all.

Page 9 and figures 6 and 7
Again, please indicate unequivocally which neurons posses which neurites, and clearly separate them in the schematic drawings (applies to all figures of this kind). In the text you write about a VUM neuron, but no primary neurite or bifurcation point is shown.

Page 14, first chapter
There is a lot of work in larger insects in which the differences between the three pterothoracic segments are described. In brief, t1 controls front legs (with mostly sensing and exploring functions) and head muscles, it also controls grooming. There are also no wings on t1. T2 and t3 are either wing bearing segments or control wings and specific structures like halteres.

Page 19
Descending octopaminergic neurons of the suboesophageal ganglion with wide spread ramifications in all segmental ganglia have been described in the locust by Bräunig and Burrows.

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N.B. These are the comments made by the referee when reviewing an earlier version of this paper. Prior to publication the manuscript has been revised in light of these comments and to address other editorial requirements.