ANAT 322 Lecture Notes - Lecture 20: Zygosity, Prolactin, Adrenocorticotropic Hormone
7 Jun 2018
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ANAT 322 Winter 2017
Lectures
Lecture 20:
→ Outputs from the Circadian Clock
2. CLOCK is always expressed whereas Bmal1 is more or less expressed depending on the time of the
day.
Clock-controlled genes is one of the molecular explanations for the outputs from the clock. CLOCK
genes were the molecular components of the clock and if we do mutations in these, it alters the
rhythms. Clock-controlled genes are controlled by the clocks but they are not part of the clock and their
RNA and proteins will be rhythmic. These could encode many different types of molecules such as other
transcription factors, neuropeptides, receptors, ion channels, metabolic enzymes and many others.
It works by CLOCK and Bmal1 binding to the CLOCK genes and other genes.
3. AVP is a small peptide involved in different physiological systems and is expressed in different parts of
the hypothalamus which includes the SCN and the supraoptic nucleus (SON) also expresses the AVP
gene and this shows in situ hybridization for AVP gene.
In the graph, the rhythm is over a 24-hour day and AVP RNA goes up during the day and down during
the night. This is reminiscent of the per1 CLOCK gene expression which is high during the day and low
during the night and it is also regulated by the same transcription factors CLOCK and Bmal1. In AVP gene
there is a DNA sequence that is the preferred sequence to be recognized by these transcription factors.
It has been found that CLOCK and Bmal1 bind to AVP gene and activate it for transcription which occurs
during the day and in the night it is repressed because the repressor is more present in the cell.
The dotted line shows CLOCK mutant mice that have a mutation in the transcription factor CLOCK and
have very low activity. If you have no activity of these transcription factors then AVP gene is very low so
you see no rhythm. However, if you look at the SON, it is always high and not affected by the CLOCK
mutation. CLOCK and Bmal1 will lead to a rhythm of AVP RNA in the SCN but not in the SON. This tell us
that CLOCK control genes are not rhythmic in all tissues but the rhythmic regulation is specific on the
tissue.
4. Here they look at this on a broader basis by using large scale transcriptomic approaches such as
microarray analysis. Researchers took SCN and liver at different times of the day and night over two
days ad they looked at thousads of gees y usig CHIPS ad hyridizig RNA’s. They ated to ko
whether these were expressed high or low depending on the time of the day.
Each line is a gene and we can see that there are many genes that present 24-hour rhythms in both
the SCN and the liver. Hundreds of gene show rhythm and different levels of RNA depending on when
we take the tissue. If we compared the genes that are rhythmic, most of them are rhythmic in either of
the tissues and only a small fraction of these are rhythmic in both tissues.
In the liver, about 20% of the transcripts are rhythmic so more than three thousand RNAs are
rhythmic. A large proportion of the transcripts are also rhythmic in many other tissues. About half of all
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ANAT 322 Winter 2017
Lectures
the genes that exist in the body are rhythmic somewhere in the body so at the molecular level, cells and
tissues do not work the same depending on the time of the day.
5. In addition to the SCN, you have clocks in other tissues or brain regions and in most peripheral organs.
If you use bioluminescence reporter mice you can put various different organs in the dish and
monitor bioluminescence. In the SCN, over four days, the bioluminescence goes up and down and this is
also seen in muscles and lungs. These tissue on their own have an endogenous clock that drives these
24-hour rhythms.
6. We have a central clock in the SCN and we have peripheral clocks in most of the other organs. The
central clock acts like the director of an orchestra. If you destroy the SCN, the different tissues will
otiue to hae rhyth ut ill hage their phase or o’t e i sy. Peripheral loks ill also hae
their own output physiological mechanisms that they control.
There are humoral cues that go through the blood stream such as cortisol or corticosterone in
rodents is one of the molecules that can allow the central clock of the SCN to reset peripheral clocks.
There are also neuronal signals such as the autonomic nervous system that allows the SCN to reset some
peripheral clocks. There are systemic cues such as the feeding rhythms and the body temperature
rhythms so the central clock tells the brain when to stimulate feeding and temperature changes. When
we have jetlag it is good to eat at the time of the country we are in because eating at regular times will
help reset the peripheral clocks.
7. The SCN will send projections to different parts of the brain such as the hypothalamus and the
thalamus that will indirectly lead to many physiological rhythms including corticosterone (cortisol in
humans), LH and melatonin rhythms.
9. Melatonin is a hormone that is synthetized by a gland called the pineal gland in the night but not
during the day and this relies on the SCN. If you destroy the SCN in rodents, you lose the melatonin
rhythm so there are connections between the two areas through complex pathways (4 different steps
between the two regions).
The pathway ends by projections from the supracervical ganglia (SCG) which are noradrenergic
projections so noradrenaline is released in the pineal gland during the night only which leads to the
rhythm of melatonin. Melatonin is synthetized from serotonin which is itself synthetized from
tryptophan. Melatonin is made from serotonin through two enzymatic steps, one involves the AA-NAT
and another through HIOMT.
AA-NAT is more important in terms of rhythmicity and it is very high in the night and very low in the
day. There is a strong circadian regulation of AA-NAT which is the limiting enzyme of the pathway. You
have a rhythm of melatonin probably because you have a rhythm of AA-NAT and this is because of the
regulation by noradrenaline. In the night, there will be more transcription of the gen that give rise to AA-
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Document Summary
Outputs from the circadian clock: clock is always expressed whereas bmal1 is more or less expressed depending on the time of the day. Clock-controlled genes is one of the molecular explanations for the outputs from the clock. Clock genes were the molecular components of the clock and if we do mutations in these, it alters the rhythms. Clock-controlled genes are controlled by the clocks but they are not part of the clock and their. These could encode many different types of molecules such as other transcription factors, neuropeptides, receptors, ion channels, metabolic enzymes and many others. In the graph, the rhythm is over a 24-hour day and avp rna goes up during the day and down during the night. This is reminiscent of the per1 clock gene expression which is high during the day and low during the night and it is also regulated by the same transcription factors clock and bmal1.
