Netrin/DCC Signaling Guides Olfactory Sensory Axons to Their Correct Location in the Olfactory Bulb

Abstract

Olfactory sensory neurons expressing particular olfactory receptors project to specific reproducible locations within the bulb. The axonal guidance cues that organize this precise projection pattern are only beginning to be identified. To aid in their identification and characterization, we generated a transgenic zebrafish line, OR111-7:IRES:Gal4, in which a small subset of olfactory sensory neurons is labeled. Most sensory neurons expressing the OR111-7 transgene project to a specific location within the bulb, the central zone protoglomerulus, while a smaller number project to the LG1 protoglomerulus. Inhibiting netrin/DCC signaling perturbs the ability of OR111-7 expressing axons to enter the olfactory bulb and alters their patterns of termination within the bulb. The netrin receptor DCC is expressed in olfactory sensory neurons around the time that they elaborate their axons, netrin1a is expressed near the medial-most margin of the olfactory bulb, and netrin1b is expressed within the ventral region of the bulb. Loss of netrin/DCC signaling components causes some OR111-7 expressing sensory axons to wander posteriorly after exiting the olfactory pit, away from netrin expressing areas in the bulb. OR111-7 expressing axons that enter the bulb target the central zone less precisely than normal, spreading away from netrin expressing regions. These pathfinding errors can be corrected by the re-expression of DCC within OR111-7 transgene expressing neurons in DCC morphant embryos. These findings implicate netrins as the only known attractants for olfactory sensory neurons, first drawing OR111-7 expressing axons into the bulb and then into the ventromedially positioned central zone protoglomerulus.

Experience-Dependent versus Experience-Independent Postembryonic Development of Distinct Groups of Zebrafish Olfactory Glomeruli

“…Our findings are consistent with previous descriptions of glomeruli in zebrafish embryos (Dynes and Ngai, 1998;Li et al, 2005;Koide et al, 2009), larvae (Li et al, 2005, and adults (Baier and Korsching, 1994;Braubach et al, 2012). Furthermore, we identified distinct maturation mechanisms among zebrafish glomeruli.…”

“…This targeted axonal migration is dependent on the presence of pioneer axons (Whitlock and Westerfield, 1998), which require functional Slit/Robo2 and chemokine signaling (Miyasaka et al, 2007) to fasciculate properly and extend into the OB primordia. Compromising the expression of Slit/Robo and other guidance molecules, such as anosmin-1 and netrin/DCC (Yanicostas et al, 2009;Madelaine et al, 2011;Lakhina et al, 2012), perturbs OSN axon targeting and leads to the formation of disorganized and incomplete glomerular arrays. Interestingly, the Robo2 mutant astray (Miyasaka et al, 2005) appears to display more severe developmental deficits in some glomeruli (lG 4 ) than in Figure 7.…”

“…Such analyses might be complicated by large increases in OSN numbers, but it is also possible that new glomeruli form de novo from new populations of OSN axons. This possibility may be addressed with zebrafish that permit visualization of individual OSN axon types (Lakhina et al, 2012), and similar strategies have already proven valuable in studying rodent OB development (Potter et al, 2001;Feinstein and Mombaerts, 2004;Zou et al, 2004). To summarize, the zebrafish glomerular map develops in two stages: (1) a coarse positional map that includes early emerging large glomeruli forms during embryonic development; and (2) this coarse map is progressively refined during 5 weeks of larval development to produce an array of discrete glomeruli that closely resemble their adult counterparts in number and distribution.…”

“…Ingrowing OSN axons follow these processes and, beginning at ϳ48 hpf, contact postsynaptic targets to form rudimentary glomeruli (Whitlock and Westerfield, 1998;Miyasaka et al, 2007). Most glomeruli appear to differentiate during larval development (Li et al, 2005), and their numbers increase more than 10-fold between prehatching embryos (Dynes and Ngai, 1998) and adults (Braubach et al, 2012). The olfactory system of adult zebrafish contains ϳ140 glomeruli, arranged in distinct groups that are innervated by specific OSN classes (Braubach et al, 2012).…”

Robo2–Slit and Dcc–Netrin1 Coordinate Neuron Axonal Pathfinding within the Embryonic Axon Tracts

“…In zebrafish, inhibition of Netrin or Dcc functions causes some olfactory sensory axons to project aberrantly and away from netrin-expressing regions (Lakhina et al, 2012). Previous studies in knock-out mice showed that DCC is required for the guidance of commissural axons to the midline (Fazeli et al, 1997).…”