Lecture 6- Clocks in The Limbic System

Question 1

What did the development of bioluminescence reporters for clock genes and proteins enable?

Answer 1

The assessment of extra SCN oscillators and in vitro clock function

Question 2

How is a bioluminescence recording prepared and measured? (wrt. clocks)

Answer 2

Sample prep: clock gene driver luciferase model
Culture: coronal/ sagittal/ horizontal angled brain slices ~250μm thick
Imaging using Luminoview system with EM-CCD camera (system at -90 degrees)

Question 3

Extra SCN oscillations in

Answer 3

OB, VLPO (arcuate nucleus, SCN, pituitary, pineal)

Question 4

Lack of rhythms in

Answer 4

Substantia nigra, piriform cortex

Question 5

Key limbic areas wrt. clocks

Answer 5

BNSTov and CeA (Am)

Question 6

How were oscillations in the BNSTov and CeA investigated?

Answer 6

Immunohistochemistry with ABs against PER2 –> in vivo assessment of clock function

Question 7

BNSTov rhythms wrt. SCN

Answer 7

Closely synchronised as strongest staining at same ZT, SCN oscillations are stronger, BNSTov depends on SCN

Question 8

SCN lesion wrt. BNSTov

Answer 8

Causes ipsilateral BNSTov to become arrhythmic

Question 9

BNSTov relationship with adrenal glands

Answer 9

PER2 expression arrhythmic when adrenal glands are ablated (BNSTov rhythms need SCN and corticosterone)

Question 10

CeA also…

Answer 10

not autonomous oscillator, requires SCN and adrenal input

Question 11

Epithalamus

Answer 11

Just above thalamus

Connects limbic system to rest of brain, contains Hb, habenula commissure, stria medullaris, pineal gland

Question 12

Hb (Habenula)

Answer 12

Relay linking limbic forebrain inputs

Roles in reproduction, nutrition, spatial memory, -ve reward, addiction, anxiety, stress, SLEEP WAKE cycles

Question 13

Relevant innervation of Hb by?

Answer 13

Melanopsin containing RGCs

Question 14

In mouse, neurons of the lateral Hb respond to…

Answer 14

…retinal illumination

Question 15

Molecular (Per1-luc) rhythms in the lateral Hb depend on…

Answer 15

…a functional molecular clock

Question 16

Like SCN, mouse lateral Hb shows…

Answer 16

…rhythms in electrical activity, but unlike SCN these depend on a functional molecular oscillator

Question 17

Lateral Hb and PK2 influence

Answer 17

Lateral Hb neurons have been shown to respond to PK2

Question 18

Cry KOs in Hb

Answer 18

remove oscillations

Question 19

Nature of the Hb oscillator

Answer 19

Semi-autonomous circadian oscillator with rhythmic clock genes and electrical activity

Question 20

MBH (mediobasal hypothalamus)

Answer 20

Arcuate, dorsomedial and ventromedial nuclei, median eminence
Main internal regulator of homeostasis

Question 21

MBH and Per2:luc

Answer 21

Shows clear oscillations which do not remain synchronised when kept as single cells out of the whole tissue environment

Question 22

Area with SCN like properties

Answer 22

dArc > DMHc > LHb > CeA

Question 23

Oscillations in single cells of

Answer 23

dArc. DMHc, LHb

Question 24

Master oscillators

Answer 24

SCN, OB, retina

Question 25

Semi-autonomous oscillators

Answer 25

LHb, DMH, ARC

Question 26

Slave oscillators

Answer 26

BNSTov, Am

Question 27

Peripheral tissues expressing clock genes

Answer 27

Liver, heart, lungs, spleen, adrenal gland, pituitary gland, cartilage