Floor Plate Issues Diversity – Non-neuronal cells – Neurons Developmental origins Midbrain dopaminergic cells Comparison to insect midline cells.

Floor Plate

Issues• Diversity

– Non-neuronal cells– Neurons

• Developmental origins• Midbrain dopaminergic cells• Comparison to insect midline cells

Spinal Cord Development

Notochord Induction of Floor Plate Formation in Mammals and Chicks

Shh

Floor Plate: Shh Induction of Spinal Cord Development

• Shh Induction of Spinal Cord Development– Patterns ventral spinal cord– v3, MN, v2, v1, v0

Floor Plate: Control of Axon Crossing

• Control of axon crossing– Netrins– Shh

Zebrafish Floor Plate Description

• Early description (Kuwada 1990)– Consists of 3 rows of cells– Defined by cuboidal shape, and most extend no processes (non-neuronal)– Among earliest identifiable cells in spinal cord and appear before first growth cones– Crossing growth cones are between FP and basal lamina next to notochord

Zebrafish Floor Plate Description

• Two cell populations (Odenthal 2000)– Medial FP (MLP): shh+, netrin1+, fkd4+, fkd7+

– Lateral FP (LFP): foxa2 (fkh)+, fkd4+, netrin-1+

Zebrafish MFP Genetics

• MFP– cyclops (cyc; nodal-TGF-), one-eyed pinhead (oep; EGF-CFC), schmalspur (sur; foxh1) result in

absence of MFP but not LFP• LFP does not require MFP for development

Zebrafish LFP Genetics

• LFP– no-tail (ntl; fly brachyenteron), floating head (flh), momo (mom) have an absence of

notochord• flh or mom: MFP present with associated LFP; LFP absent when MFP absent• In ntl: get expansion of MFP and LFP• LFP can develop without notochord (as can MFP), but LFP only in presence of MFP

Shh Control of LFP

• syu (sonic you: shh)– Affects LFP but not MFP– Inject syu RNA increase LFP but not MFP

Zebrafish Summary

• Early cells decide to be either notochord or FP• MFP: notochord and shh-independent; requires nodal-related pathway (TGF-)• LFP: dependent on shh from either Notochord or MFP• Species differences:

– Murine and chicken shh from notochord is required for induction of all FP cells– Murine Shh and netrin-1 are only in MFP and foxa2 (fly fkh) broadly expressed– Chicken Foxa2 in MFP and shh and netrin-1 in LFP

Multiple Floor Plate Cell types

• Zebrafish (Schäfer 2007)– foxa2: all MFP; subset LFP– tal2 bHLH gene (fly ~HLH4C) in subset of foxa2– LFP

• Insect comparison– Both insect midline and vertebrate FP consist of neurons and non-neuronal (glial) cell types

Tal2+ LFP cells are Postmitotic

• Label with pHH3– Tal2+ cells negative– Foxa2 + tal2– cells positive

• Tal2 + cells are postmitotic

Multiple Floor Plate Cell types

• Late stage embryonic FP– Sim1+ cells are LFP subset and postmitotic– Sim1+ cells express neuronal marker– Sim1+ cells are V3 interneurons (glutamatergic in other contexts)– Also observed GABAergic neurons

Summary

• Tal2 + cells are neuronal progenitor cells– Subset sim1 + V3 interneurons– May also give rise to other interneurons

• Tal2 – foxa2+ cells are non-neuronal• Insect comparison

– Both insect ML and vertebrate have Sim + interneurons

Notch Signaling

• Mindbomb (Neur) mutant– Nkx2.2b (vnd), foxa2 (LFP non-neuronal cells), tal2 (LFP neurons) reduced or absent– Foxa2 in MFP (non-neuronal) normal

• Notch signaling required for LFP and p3 neurons

wt

wt

mib

mib

Shh Signaling

• In shh mutants, p3 LFP cells absent• In dtr/+ and yot/+ foxa2 LFP was absent and tal2 and sim1 normal• Thus, non-neuronal LFP cells require high Shh levels compared to p3 cells

Summary

• Early– foxa2 + cells proliferate– Tal2 + cells are postmitotic and differentiate into neurons

• Later– Subset of tal2+ cells become sim1 + V3 interneurons– Also GABAergic KA neurons (sim1 +?)

• Notch signaling required for LFP and p3 cells• Shh signaling: LFP non-neuronal and neuronal cells required different levels of Shh

Midbrain FP Cells

• Midbrain dopaminergic neurons– Motor control, reward, addiction, attention, cognition– Loss results in Parkinson disease

• Origins: mouse (Kittappa 2007)– TH+ cells are in foxa2+ FP cells in midbrain

Midbrain FP Cells

• Shh is expressed in FP• Shh-cre X R26R lacZ reporter: lineage marker• TH+ cells overlap with -Gal

Development of Midbrain FP Cells

Dorsal fate

5-HT1a fate

Midbrain FP Cells

• Progenitor cell development– Shh: induces FP– Otx2: required for generation of mesDA precursors

– Lmx1a (CG32105): required for mesDA cell fate– Ngn2: proneural function

• Postmitotic development– Nurr1: controls subset of mesDA gene expression (TH)– Lmx1b (CG4328): survival; affects Pitx3 but not other

genes (Nurr1, early TH)– Pitx3: lose TH expression, affects migration, survival– En: required for late maturation and survival

Foxa2

• Kittappa 2007• Foxa2–/–

– No TH+ cells

• Foxa2+/–

– Motor defects– Loss of DA neurons– Like PD

En

• En (Simon 2004)• En is gradually expressed in TH + cells• En –/–

– Early TH+ cells present but lack fibers– Late TH+ cells absent

Summary

• Vertebrate FP:insect ML– Diversity of neuronal and non-neuronal cell types– MG = non-neuronal FP; both important signaling centers– Sim+ cells present– DA+ cells present– Genes?