Songbirds, Steroids, and Adult

Brad Walters Biological Sciences Lehigh University September, 2008 Songbirds, steroids, and neurogenesis… Some tissues regenerate well…

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http://flickr.com/photos/museumoflondon/2392340925/ http://www.anat.ucl.ac.uk/business/becker1.shtml Others, not so good…

http://commons.wikimedia.org/wiki/Image:MCAO-sheep.jpg

The absence of evidence is not evidence of absence!!! Visualizing Neurogenesis: (e.g. ) get incorporated into DNA strands during replication

Altman, 1962

[3H] Thymidine incorporates into dividing cells!

http://www.bcscience.com/bc9/pgs/quiz_section5.1.htm Some songbird brains change dramatically across seasons

Brenowitz et al, 1991

Margoliash, 2005

Nottebohm, 1981 Nucleus Do the changes in Hillock volume result from Neurogenesis?

Axon Dendrites

A stained with [3H]thymidine, counterstained with methylene blue. Bar = 50µm Electron micrograph of a new neuron in HVC. N = nucleus, A = axonal hillock, Goldman and Nottebohm, 1983 D = dendrite,

But are they really ? Voltage (mV) Voltage Time (msec)

Paton and Nottebohm, 1984 No… But seriously… Are they really neurons?

The retrograde tracer, Fluorogold was injected into RA, a nucleus known to be innervated by projecting from HVC.

Kirn and Nottebohm, 1991

Seasonal neurogenesis in songbirds suggests the involvement of steroid hormones. Spring

Fall

Kirn and Nottebohm, 1993 Smith et al. 1997 increases neurogenesis

Bottjer and Dignan, 1988 However, Testosterone can be metabolized in the brain…

5-α Reductase Aromatase

Testosterone

Dihydrotestosterone (DHT) 17β-Estradiol (Estrogen) So which steroid is it?

C T DHT E2 DHT+E2 C T DHT E2 DHT+E2

Tramontin et al. 2003 Estrogen increases neurogenesis in adult songbirds…

Average # of HVC Average # of New HVC Neurons/Section Neurons/Section 18

200 16

* 14 *

150 12

10

100 8 HVC Neurons 6

50 [3H] Positive HVC Neurons N = 7 N = 8 4 N = 7 N = 8 2

0 0 No12 E E Replaced No12 E E Replaced

adapted from Hidalgo et al. 1995 Estrogens enhance mammalian neurogenesis.

Tanapat et al. 1999 Proliferation Migration

Integration

Perpetuation

Differentiation

Adapted from Gage, 2003 Which process or processes are being affected by Estrogen provision?

• Functional 1. Proliferation Neurogenesis is 2. Migration comprised of… 3. Differentiation 4. Perpetuation (Survival) 5. Integration Does E affect neural stem proliferation? Two main questions: – Do estrogens increase the number of cells entering the cell cycle at any given point in time?

– Do estrogens increase the rate at which cells proceed through the cell cycle? Does E affect neuronal migration?

• E acts on cells involved in the migration process. • E influences migration away from VZ.

Williams et al., 1999 Does E affect Differentiation?

• Does E affect cell fate directly? • Does E affect factors

that affect cell fate? 200 BMP-2

150

100

50 Fold Change of mRNA expression Fold of mRNA Change

0 Brànnvall et al. 2002 Fadrozole Saline Modified from Walters and Saldanha, 2008 Perpetuation (Survival)

**

FAD FAD+E2

Saldanha et al. 2005 With regard to adult neurogenesis, estrogen affects…

• Proliferation: tentative yes • Migration: tentative yes • Differentiation: maybe • Survival: yes • Integration: ??? This is the next big question!!! OTHER FACTORS KNOWN TO INFLUENCE NEUROGENESIS INCLUDE: HORMONES (-) NEUROTRANSMITTERS (+) Testosterone (+) Norepinephrine (+) Progesterone (+) Glutamate (-)

BEHAVIOR (+) Enriched Environment (+) Gage, 2003 Summary

• Brain Injury and Neurodegenerative diseases can be devastating to the adult brain. • However, neurogenesis in the CNS does occur throughout life. • Thymidine analogs are a powerful, but limited, tool for measuring this process. • Songbirds are an indispensable in this field. • Steroid hormones affect neurogenesis. • Estrogen increases neurogenesis most likely by increasing survival as well as via proliferation (probably), and possibly by affecting cell fate and/or migration. • There are many factors, often working together, that affect neurogenesis. • Clinical applications will depend heavily on the site specific migration and integration of new neurons. Acknowledgements

PI Colin Saldanha

Collaborators Ryan Wynne

Work funded by: NIH (NINDS) 047267