DEVELOPMENT OF THE OLFACTORY BULB
OVERVIEW
The sense of smell in rats is one of the most dominant in the regulation of their behavior. Smells in their environments allows them to construct maps to find food, mates, avoid dangerous places, and seek safe places to live and care for their offspring. Rats are an extremely adaptable species and are successful because they can live in environments that are disturbed by human activity—all that flexibility in their behavior is largely due to the sense of smell.
The olfactory bulb itself is a unique “outpost” in the forebrain. Every cell population in the olfactory bulb migrates into the bulb from germinal sources in the basal forebrain. The mitral cells of the MOB leave their axons behind in the lateral olfactory tract (LOT) as they migrate forward to create the protrusion known as the olfactory bulb. The impetus for mitral cell migration is probably due to a chemical attractant present in the olfactory nerve fibers that contact the forebrain before there is any discernible olfactory bulb. The LOT is one of the major fiber tracts below the rhinal sulcus that distributes olfactory sensory information to the anterior olfactory nucleus, olfactory cortex, amygdala, entorhinal cortex, and field CA1 of the ventral hippocampus without a relay station in the thalamus. New short-axon interneurons are constantly being generated by stem cells in the rostral migratory stream–a conduit of young neurons generated by neuronal precursors in the subependymal layer of the anterior horns of the lateral ventricles. The widely-distributed olfactory sensory information has a profound effect on the behavioral responses chosen to adapt to environmental changes.
This project is a series of 142 images that are labeled, annotated, and “zoomified” so that users can explore the complete prenatal developmental sequence of the main olfactory bulb (MOB) and the accessory olfactory bulb (AOB). By clicking on a thumbnail in an image set, a new window will appear that displays the high resolution file. A user can view low to high magnification to see the entire slice or “zoom” to small areas where individual cells and fibers can be examined; each image has an embedded navigator to quickly select the area for high-magnification viewing. The 10 image sets that follow are arranged according to age from early (Ell, E12, and E13) to late (E22, the day before birth). For each age group except E22, 3H‑thymidine autoradiograms are mixed with methacrylate-embedded normal embryos. In this way, the labeling patterns in the autoradiograms can inform us about the identity of migrating and settling cells in the olfactory bulb on a specific embryonic day. The thumbnails in each image set have different shapes because of the variation in the section planes. Text above the rows of thumbnails give some brief information about the contents of each slice.
At the bottom of each image set and this overview, there are Jumps inserted so that users can examine individual images from different age groups. For example, suppose one wants to look at the ventral horizontal methacrylate slice from each image set; users can jump from one age group to another to display the horizontal images they want to examine–each will open in a new page–then easily see changes from early-to-late prenatal development. It is also interesting to view the early-to-late P5 survival series as a separate group of images. There are endless possibilities for use of these images. Remember, if you copy these images into your own publications, please give credit to this website.
The research used to identify structures in the olfactory bulb comes from previous publications of members of the Laboratory of Developmental Neurobiology. You can find pdfs of the original articles here: http://neurondevelopment.org/olfactory-development/
Thank you!
Shirley A. Bayer
August 22, 2017
What happens in the developing olfactory bulb on Embryonic Days (E)11, 12, and 13?
THERE ARE 11 IMAGES IN THIS SET; CLICK ON THE THUMBNAILS BELOW TO VIEW HIGH RESOLUTION IMAGES.
Summary of development in the earliest age groups:
First, there are very few neurons produced on E11 and E12, and there is no olfactory bulb extending from the basal forebrain on all 3 days. Instead, the germinal sources of the projection neurons in both the accessory olfactory bulb (AOB) and the main olfactory bulb (MOB) are actively proliferating and adding more precursor cells to their populations in the neuroepithelium (NEP) of the basal forebrain–the stockbuilding phase of population development. During the day on E13, 60% of the AOB output neurons are being generated–the production phase of population development.
Changes in the skull interact with what is happening in the brain and ultimately influence the brain to produce an olfactory bulb. The chief skull change is that the flattened olfactory placodes on the lateral edges of the skull on E11 grow and move inward on E12 and E13 (invaginate) to create the primitive nasal cavity anterolateral to the basal forebrain. A portion of the invaginating cells will become the olfactory epithelium that will house the primary sensory olfactory neurons; the axons of these neurons form the olfactory nerve that grows toward the most anterior extent of the forebrain. The olfactory nerve does not actually contact the forebrain on E11, E12, or E13, but the nerve is much longer and closer to the brain on E13 than it is on E12. With regard to the AOB, a portion of the nasal cavity will differentiate into Jacobson’s organ that is surrounded by a semicircular skull bone, the vomer. Jacobson’s organ is the source of the vomeronasal nerve that contacts the AOB.
2 consecutive 3H‑thymidine injections starting on E11, E12, and E13 and survival to E21 (Figs. 1-3).
2 consecutive 3H‑thymidine injections starting on E13 and survival to Postnatal day (P) 5 (Figs. 4-5).
24 hr survival after a 3H‑thymidine injection on E12 (Fig. 6); 2 hr survival after a 3H‑thymidine injection on E13 (Fig. 7).
Methacrylate-embedded E13 embryos in the sagittal plane (Figs. 8-9).
Methacrylate-embedded E13 embryos in the horizontal (Fig. 10), and coronal planes (Fig. 11).
What happens in the developing olfactory bulb on Embryonic Day 14?
FIGURES 12-24
By the morning of E14, nearly 70% of the AOB output neurons, 10-12% of the MOB mitral, internal tufted, and external tufted cells have already been generated. From the morning of E14 to the morning of E15, another 20% of the AOB output neurons and another 10-15% of the MOB mitral and internal tufted cells will be added. The germinal source of AOB output neurons in NEP is very thick with many postmitotic neurons “sequestered” in the superficial parts of the layer. But, major cell migration has not yet begun on the morning of E14.
The invagination of the olfactory epithelium continues on E14 and becomes well-established in the superior nasal cavity. The same is true for Jacobson’s organ that forms a hook-shaped structure in the ventromedial nasal cavity. Many more nerve fibers are growing from both the olfactory epithelium and Jacobson’s organ toward the brain. The olfactory nerve appears to make contact with the anteroventral forebrain. However, there is still no protrusion that can be recognized as an olfactory bulb.
3H‑thymidine injections starting on E14 and a subsequent injection on E15, survival to E21 (Fig. 12)
3H‑thymidine injections starting on E14 and a subsequent injection on E15 with survival to Postnatal day (P)5 (Figs. 13-14).
The 24 hr (Fig. 15) and 2hr (Fig. 16) survival specimens after single 3H‑thymidine injections on E13 and E14.
High resolution images of the methacrylate-embedded E14 embryos in the sagittal plane (Figs. 17-19).
High resolution images of the methacrylate-embedded E14 embryos in the horizontal plane (Figs. 20-22).
High resolution images of the methacrylate-embedded E14 embryos in the coronal plane (Figs. 23-24).
What happens in the developing olfactory bulb on Embryonic Day 15?
FIGURES 25-40
The proportion of neuronal populations generated before the morning of E15: 89%-AOB output neurons; 22%-MOB mitral cells; 25%-MOB internal tufted cells; 10%-external tufted cells.
The proportions that will be added on E15 and by the morning of E16: 5%-AOB output neurons; 47%-MOB mitral cells; 12%-MOB internal tufted cells; 1%-external tufted cells.
The germinal source of the AOB output neurons is tentatively identified in a horizontal methacrylate slice (Fig. 36) and cells can be traced migrating forward toward the anterior forebrain, but there is not a distinguishable AOB. Tentatively identified mitral cells can be seen migrating in other horizontal slices (Figs. 37-38). Indeed, the olfactory bulb itself is very indistinct, but there is a large part of the basal forebrain that is contacted by the olfactory nerve. The vomeronasal nerve contacts the dorsomedial part of the olfactory part of the basal forebrain and may attract AOB output neurons to migrate in that direction. So E15 is the beginning of the appearance of migratory streams toward the olfactory bulb.
3H‑thymidine injections on E15 and E16, survival to E21 (Fig. 25)
3H‑thymidine injections starting on E15 and E16, survival to Postnatal day (P)5 (Figs. 26-27).
The 24 hr (Figs. 28-29) survival specimens after a single 3H‑thymidine injection on E14.
The 2hr (Figs. 30-32) survival specimens after a single 3H‑thymidine injection on E15.
High resolution images of a methacrylate-embedded E15 embryo in the sagittal plane (Figs. 33-35).
High resolution images of a methacrylate-embedded E15 embryo in the horizontal plane (Figs. 36-38).
High resolution images of a methacrylate-embedded E15 embryo in the coronal plane (Figs. 39-40).
What happens in the developing olfactory bulb on Embryonic Day 16?
FIGURES 41-56
The proportion of neuronal populations generated before the morning of E16: 94%-AOB output neurons; 69%-MOB mitral cells; 37%-MOB internal tufted cells; 11%-external tufted cells.
The proportions that will be added on E16 and by the morning of E17: 6%-AOB output neurons; 25%-MOB mitral cells; 22%-MOB internal tufted cells; 15%-external tufted cells.
The germinal source of the AOB output neurons is more definitely identified in a dorsal horizontal methacrylate slice (Fig. 51) and cells can be traced migrating forward toward the anterior forebrain, but there is still not a distinguishable AOB. Migrating mitral cells are prominent in the other horizontal slices (Figs. 53-55) and 2 different streams (lateral and medial) are shown. But the mitral cell layer in the olfactory bulb is not there yet. The cells in the olfactory bulb, which is finally a definite protrusion surrounded by the olfactory and vomeronasal nerves have no laminar differentiation. E16 heralds the beginning of migratiory paths into the bulb.
3H‑thymidine injections starting on E16 and a subsequent injection on E17, survivals to E21 (Fig. 41).
3H‑thymidine injections starting on E16 and a subsequent injection on E17, survival to Postnatal day (P)5 (Figs. 42-43).
24 hr survival after a single 3H‑thymidine injection on E15 (Figs. 44-45)
2hr survival after a 3H‑thymidine injection on E16 (Figs. 46-48).
High resolution images of methacrylate-embedded E16 embryos in the sagittal plane (Figs. 49-50).
High resolution images of methacrylate-embedded E16 embryos in the horizontal plane (Figs. 51-54).
High resolution images of methacrylate-embedded E16 embryos in the coronal plane (Figs. 55-56).
What happens in the developing olfactory bulb on Embryonic Day 17?
FIGURES 57-71
The proportion of neuronal populations generated before the morning of E17: 100%-AOB output neurons; 94%-MOB mitral cells; 59%-MOB internal tufted cells; 26%-external tufted cells.
The proportions that will be added on E17 and by the morning of E18: 5%-MOB mitral cells; 32%-MOB internal tufted cells; 12%-external tufted cells; 5%-interstitial tufted cells.
The migratory stream of the AOB output neurons is still visible, but not as robust as on E16. In contrast, the two mitral cell migratory streams are becoming more prominent, and more axons are accumulating in the lateral olfactory tract. There is still very little laminar differentiation inside the growing olfactory bulb because of the massive numbers of cells migrating into it from germinal sources in the basal telencephalon.
3H‑thymidine injections on E17 and E18, survival to E21 (Fig. 57).
3H‑thymidine injections on E17 and E18 with survival to Postnatal day (P)5 (Figs. 58-59).
24 hr survival after a single 3H‑thymidine injection on E16 (Figs. 60-62).
2 hr survival after a single 3H‑thymidine injection on E17 (Figs. 63-64).
High resolution images of a methacrylate-embedded E17 embryo in the sagittal plane (Figs. 65-66).
High resolution images of a methacrylate-embedded E17 embryo in the horizontal plane (Figs. 67-69).
High resolution images of a methacrylate-embedded E17 embryo in the sagittal coronal plane (Figs. 70-71).
What happens in the developing olfactory bulb on Embryonic Day 18?
FIGURES 72-88
The proportion of neuronal populations generated before the morning of E18: 100%-AOB output neurons; 99%-MOB mitral cells; 91%-MOB internal tufted cells; 38%-external tufted cells; 5%-interstitial tufted cells..
The proportions that will be added on E18 and by the morning of E19: 1%-MOB mitral cells; 7%-MOB internal tufted cells; 27%-external tufted cells; 11%-interstitial tufted cells.
The olfactory bulb continues to grow and is now one of the most prominent structures in the basal forebrain. For the first time there is a true laminar differentiation in the olfactory bulb, and an AOB can be distinguished in the posterodorsal region. The mitral cell migratory streams are very prominent and the cells can be seen streaming out of the enlarging lateral olfactory tract.
3H‑thymidine consecutive exposures on E18 and E19, survival to Postnatal day (P)5 (Figs. 72-73).
24 hr survival after a single 3H‑thymidine exposure on E17 (Figs. 74-75).
24 hr survival after a single 3H‑thymidine exposure on E17 (Figs. 76-77).
2 hr survival after a 3H‑thymidine exposure on E18 (Figs. 78-79).
2 hr survival after a 3H‑thymidine exposure on E18 (Figs. 80-81).
High resolution images of methacrylate-embedded E18 embryos in the sagittal plane (Figs. 82-83).
High resolution images of methacrylate-embedded E18 embryos in the horizontal plane (Figs. 84-86).
High resolution images of a methacrylate-embedded E18 embryo in the coronal plane (Figs. 87-88).
What happens in the developing olfactory bulb on Embryonic Day 19?
FIGURES 89-104
The proportion of neuronal populations generated before the morning of E19: 100%-AOB output neurons; 100%-MOB mitral cells; 98%-MOB internal tufted cells; 65%-external tufted cells; 16%-interstitial tufted cells..
The proportions that will be added on E19 and by the morning of E20: 1%-MOB internal tufted cells; 29%-external tufted cells; 19%-interstitial tufted cells.
The olfactory bulb continues to grow larger. Layers in the olfactory bulb are becoming more distinct in both the AOB and the MOB; the AOB now has a recognizable granule cell layer. The mitral cell migratory streams are still very prominent and the cells can be seen streaming out of the enlarging lateral olfactory tract.
3H‑thymidine consecutive exposures on E19 and E20, survival to Postnatal day (P)5 (Figs. 89-90).
24 hr survival after a single 3H‑thymidine exposure on E18 (Figs. 91-92).
24 hr survival after a single 3H‑thymidine exposure on E18 (Figs. 93-94).
2 hr survival after a 3H‑thymidine exposure on E19 (Figs. 95-97).
High resolution images of methacrylate-embedded E19 embryos in the sagittal plane (Figs. 98-99).
High resolution images of methacrylate-embedded E19 embryos in the horizontal plane (Figs. 100-102).
High resolution images of a methacrylate-embedded E19 embryo in the coronal plane (Figs. 103-104).
What happens in the developing olfactory bulb on Embryonic Day 20?
FIGURES 105-122
The proportion of neuronal populations generated before the morning of E20: 100%-AOB output neurons; 100%-MOB mitral cells; 99%-MOB internal tufted cells; 90%-external tufted cells; 35%-interstitial tufted cells..
The proportions that will be added on E20 and by the morning of E21: 1%-MOB internal tufted cells; 7%-external tufted cells; 34%-interstitial tufted cells. Neurogenesis of the output neurons is nearly completed. Most neurogenesis is occurring in the interneuronal populations of the MOB and the rostral migratory stream is becoming established; that will persist into adulthood.
Growth in the olfactory bulb continues at a rapid pace. The AOB is now clearly recognized as a distinctly different region than the MOB. Layers in both parts are better defined; the granule cell layer in the AOB can be distinguished by fiber infiltration. The mitral cell migratory streams are still prominent in the ventral horizontal slice (Fig. 118) and mitral cells stream out of the enlarging lateral olfactory tract.
3H‑thymidine consecutive exposures on E20 and E21, survival to Postnatal day (P)5 (Figs. 105-106).
24 hr survival after a single 3H‑thymidine exposure on E19 (Figs. 107-108).
24 hr survival after a single 3H‑thymidine exposure on E19 (Figs. 109-110).
2 hr survival after a 3H‑thymidine exposure on E20 (Figs. 111-113).
High resolution images of methacrylate-embedded E20 embryos in the sagittal plane (Figs. 114-115).
High resolution images of methacrylate-embedded E20 embryos in the horizontal plane (Figs. 116-118).
High resolution images of a methacrylate-embedded E20 embryo in the horizontal-coronal plane (Figs. 119-120).
High resolution images of a methacrylate-embedded E20 embryo in the coronal plane (Figs. 121-122).
What happens in the developing olfactory bulb on Embryonic Day 21?
FIGURES 123-135
The proportion of neuronal populations generated before the morning of E20: 100%-AOB output neurons; 100%-MOB mitral cells; 100%-MOB internal tufted cells; 97%-external tufted cells; 69%-interstitial tufted cells..
The proportions that will be added on E20 and E21: 3%-external tufted cells; 29%-interstitial tufted cells.
The biggest difference from E20 to E21 is the enormous growth in the olfactory bulb as neurons differentiate and take up more space for growing axons and dendrites. The fibers in the AOB granule cell layer go in between pyramidal-shaped clumps of closely-packed cells. The outermost part of the MOB granular layer shows larger, more differentiated cells than the part next to the SVZ. Mitral cell migratory streams are still visible, but not as prominent as E20 and younger specimens.
3H‑thymidine consecutive exposures on E21 and E22, survival to Postnatal day (P)5 (Figs. 123-124).
24 hr survival after a single 3H‑thymidine exposure on E20 (sagittal, Figs. 125-25).
24 hr survival after a single 3H‑thymidine exposure on E20 (coronalal, Figs. 127-128).
High resolution images of methacrylate-embedded E21 embryos in the sagittal plane (Figs. 129-130).
High resolution images of methacrylate-embedded E21 embryos in the horizontal plane (Figs. 131-133).
High resolution images of a methacrylate-embedded E21 embryo in the coronal plane (Figs. 134-135).
What happens in the developing olfactory bulb on Embryonic Day 22?
FIGURES 136-142
There are no injection groups where 3H‑thymidine injections began on E22; a second injection would have disturbed the pregnant dams just at the time that they were giving birth or had already delivered their pups. We discuss the methacrylate-embedded E22 specimens.
The olfactory bulb grows larger as differentiation continues. The layering approaches the clarity of the P5 specimens. The big giveaway that we are looking at prenatal specimens is the oozing of cells into the lumen of the anterior olfactory recess of the lateral ventricle. Many features showing up in E21 are more distinct and prominent. There is a more clear maturation gradient between the outermost part of the MOB granular layer and the smaller neurons near the SVZ. The mitral cell migratory streams are less distinct because most of the mitral and tufted cells have already settled.
High resolution images of methacrylate-embedded E22 embryos in the sagittal plane (Figs. 136-137).
High resolution images of methacrylate-embedded E22 embryos in the horizontal plane (Figs. 138-140).
High resolution images of a methacrylate-embedded E22 embryo in the coronal plane (Figs. 141-142).
