BDNF Seminar Spring 2010 Maneeshi Prasad Jan 29th 2010

dlPFC (dorsolateral prefrontal cortex) dlPFC is the last area (45th) to develop (myelinate) in the human cerebrum. dlPFC is connected to the orbitofrontal cortex, thalamus, parts of the basal ganglia (the dorsal caudate nucleus), the hippocampus, and primary and secondary association areas of neocortex, including posterior temporal, parietal, and occipital areas.

Jon S. Simons & Hugo J. Spiers Nature Reviews Neuroscience 4, 637-648 (August 2003)

dlPFC dlPFC serves as the highest cortical area responsible for motor planning, organization, and regulation. It plays an important role in the integration of sensory and mnemonic information and the regulation of intellectual function and action. It is also involved in working memory. Complex mental activities require additional cortical and subcortical circuits that are connected with dlPFC. dlPFC development and maturation may last up to two decades in humans and may thus show differential expression of genes.

BDNF Brain-derived neurotrophic factor belongs to "neurotrophin" family of growth factors BDNF support growth and differentiation of new neurons and synapses in CNS and PNS BDNF is active in the hippocampus, cortex, cerebellum and basal forebrain—areas vital to learning, memory, and higher thinking BDNF is also involved in neurogenesis BDNF binds to TrkB receptor and p75NTR receptor

BDNF in CNS Alternative promoter usage provides differential mRNA stability and subcellular localization Promoter IV of BDNF has been implicated in forming inhibitory synapses in the cortex BDNF mRNA with long 3’UTR are localized in dendrites of cortical neurons, while short 3’UTR mRNA is restricted to soma Loss of long 3’UTR mRNA results in denser and thinner dendritic spines of CA1 pyramidal neurons and reduced hippocampal long- term potentiation Rat visual cortex shows expression of transcripts III-V and IV-V in cell soma, while IV-V is expressed in dendritic processes

BDNF in CNS Early postnatal development shows greater increase in spine density which is reduced by ~40% in later life Conversion of proBDNF to mBDNF promotes late-phase long-term potentiation expression in hippocampus ProBDNF is higher during postnatal stages while mBDNF is prominent in adults

Activity-dependent BDNF Expression Influences Homeostatic Plasticity Activity-dependent BDNF Expression Influences Homeostatic Plasticity. Excitatory neuronal activity increases postsynaptic BDNF levels via Ca2+ -dependent transcription factors. For example, CREB phosphorylated on serine 133 binds to CaRE3/CRE following coactivation by CREB binding protein (CBP). Postsynaptic release of BDNF subsequently promotes the formation of inhibitory GABAergic synapses. The BDNF precursor, pro-BDNF is also an actively secreted molecule that affects synaptic plasticity during development. Yang, J. et al. (2009) Nat. Neurosci. 12:113.

BDNF during development Plays a important role during cortical development and is required for formation of ocular dominance columns in visual cortex High levels of BDNF mRNA its TrkB receptor has been seen in dlPFC of young adults which subsequently decreases in adults

DNA-RNA-Protein Gene locus on chromosome 1 2 3 DNA Transcription mRNA 1 2 3 2 3 Translation Pre-pro Protein Protein Processing by cleavage Mature protein

Alternative transcripts of BDNF West et al, 2001

 Brain-derived neurotrophic factor (BDNF) gene structure

Human BDNF gene structure 10 noncoding exons in the 5’UTR Presence of multiple translational (ATG) start sites The Brian

Table. 1. Brain cohort demographics

Demographic variables Postmortem pH values: 6.12-6.98 Postmortem Interval (PMI): 4 hr-32 hr RNA integrity (RIN) varied

Expression of housekeeping genes across development Real time PCR principle: http://www.youtube.com/watch?v=8jBSAca9XNU No variation in mRNA expression of housekeeping genes with changes in pH and RIN

BDNF mRNA expression in dlPFC during development

BDNF mRNA expression in dlPFC during development Transcript I-IX changed significantly across development Higher during earlier stages Peaked at infancy Decreased after infancy Constant level was maintained from school age till adulthood

BDNF mRNA expression in dlPFC during development Transcript II-IX was low at birth, increased during 1st few years and peaked in toddlers Lowest in neonates Increased in infants and toddlers Decreased during school age, similar to infants, and was maintained till adulthood

BDNF mRNA expression in dlPFC during development Transcript IV-IX highest in infants and toddlers Lowest in neonates Increased in infants and toddlers age group Decreased gradually from school age and stayed consistent from adolescent till adulthood

BDNF mRNA expression in dlPFC during development Transcript VI-IX peaks within first years of life Highest at infancy Decreased subsequently from toddler age till adulthood

Real-Time PCR

v BDNF Protein levels

BDNF protein expression in dlPFC during development Both proBDNF (28 kDa) and mature BDNF (14 kDa) bands were seen at all ages Protein expression increased from neonates to infants Infants had highest level of BDNF expression followed by toddlers Mature BDNF form varied across development and peaked at infancy Increase of protein level in toddler age group might be related to increase in level of IV-IX or II-IX transcripts Decrease in protein levels in adults matches with decrease in mRNA levels Decrease in level of pre-proBDNF was similar to that of mature BDNF

BDNF distribution in dlPFC by ISH BDNF transcript

Expression of BDNF transcripts in layer IV

Expression of BDNF transcripts in layer V & VI

BDNF distribution in dlPFC All 4 BDNF transcripts were highest in deeper cortical layers V and VI Layer I: no expression at any age group Layer II: moderate expression Layer III: robust expression was seen in neonates Layer IV/mid cortical layer: lower expression neonates and low to moderate expression in older age group Layer V and VI: intense staining for BDNF seen in neurons

Discussion Transcripts I-IX, IV-IX and VI-IX had highest expression patters in infancy II-IX transcript was highest at toddler age group and was delayed by 2-3 years as compared to the other 3 transcripts Lower expression of all transcripts during school age years

Discussion DLPFC layer IV showed increased BDNF signal DLPFC layer IV is enriched in inhibitory neurons: cannot express BDNF by itself This BDNF signal may be due to the BDNF mRNA that is targeted to the apical dendrites of layer V pyramidal neurons

Discussion High level of BDNF transcripts & protein during early years and overlaps with 1.5 fold increase in synaptic density Synaptic density may decrease during adolescence and stabilize by young adulthood with cortex reaching maturity, which overlaps with the decrease in BDNF levels

Discussion Current study differs from a previous study (Webster, 2002), where BDNF expression was lowest in infancy and higher in young adult group This may be due to combining of neonatal and infant groups, and differences in cohorts

Conclusions The dynamic regulation of BDNF gene in hDLPFC may be activated in a promoter-specific manner During postnatal cortical development, neuronal morphology and synaptic density may be regulated by transcript specific BDNF expression

Thank You !