Aberrant Light Directly Impairs Mood and Learning Through Melanopsin-expressing Neurons

Legates, T., Altimus, C., Wang, H., Lee, H., Yang, S., Zhao, H., Kirkwood, A., Weber, T., & Hattar, S.

Nature, 491(7425), 594-598, November 2012

Learning Objectives• What makes intrinsically photosensitive retinal ganglion cells

(ipRGCs) “intrinsically photosensitive”?• What properties of ipRGCs make them suited for relaying

irradiance information over long periods of time?• In what ways have various subtypes of ipRGCs been

differentiated?• What evidence extends the functional roles of ipRGcs beyond

that of “non-image forming” (photoentrainment and pupillary light reflex)?

• In what ways can circadian rhythms be measured?• In what ways could aberrant light have a direct impact on

humans, despite the presence of normal sleep/circadian architecture?

Background

• ipGRCs and melanopsin

Background• What is the importance of our circadian

rhythm?• Link between sleep deprivation and

sleep/circadian disruption well established• Can irregular light directly impact mood and

cognitive functioning despite normal sleep and circadian patterns?

Methods

• Animals and Housing– Adult wild-type mice (4-8 months)– aDTA mice (6 - 8 months) • By 6 months, melanopsin cell population dies

– Both initially entrained to a 12:12LD cycle (T24)• One group switched to 3.5:3.5LD cycle (T7) for 2 weeks

– Light held constant at ~800lux• Full Daylight is about 10,000 LUX • Cloudy day is about 1,000 LUX • A lighted parking lot at night is about 10 LUX (average) • A full moon is about 0.1 LUX

• Circadian timing measures– Body Temperature • Telemetric probes

– General activity measure• Infrared motion detectors• 10 min samples

Methods (Circadian Rhythms)

• Molecular rhythm measures (PER2 & corticosterone)– Real-time polymerase chain rection (PCR)

• PCR used to detect and/or quantify target DNA molecules

– PER2 immunohistochemistry• Used to detect different protein-expressions in cells of a tissue sample

– Corticosterone measurement• Serum collected from tail blood• Differences over time, and between lighting conditions

Methods (Circadian Rhythms)

Methods (Neuronal Activity)

• Light-induced c-fos expression– indirect marker of neuronal activity – Increased c-fos mRNA can indicate recent activity– Some SCN sections were stained

immunohistochemically for c-fos• Others were incubated in rabbit

anti-c-fos, then mounted to slides– C-fos positive cells were counted and their area

measured

Methods (Depressive-behaviors)• Sucrose Anhedonia– Anhedonia describes the loss of interest– Preference measured over 2 days after

being acclimated• Forced Swim test– Immobility measured during last 4 of 6

minutes • Open Field test– Percent distance traveled and time in

center measured

Methods (Anxiety-related behaviors)

• Light-dark Box– Bright (600 lux) and dim (<1lux) areas– # of transitions b/t areas, time spent in

lit room, and distance traveled • Elevated Plus maze– Open arms (~600 lux) / Closed (~200lux)– Time spent / distance traveled in open

arms measured

Methods (Spatial Memory)• Morris Water Maze– Hippocampal-dependent task– Four stages of testing over 12 days total

• Visual – Used to familiarize mice with test– Also a screen for floating behaviors

• Acquisition– Trained 1 trial/day for 12 days to find platform– Used 4 visual distal cues surrounding the pool

• Probe– Platform removed on Day 13– Time in target quadrant measured*

• Reversal– Platform moved to opposite quadrant on Day 14– Mice trained as done in first acquisition phase

Methods (Recognition memory)• Previous testing ensured • 3 Stages– Acclimation to arena– Familiar object exposure• 10 min, then returned to home

– Novel object exposure• After 1 hour, placed back in arena (5 min)

– Percent of time spent w/ each object measured

Methods (Long-term potentiation)• Coronal hippocampal slices • Synaptic responses evoked after 20

min baseline – LTP induced by theta burst stimulation

(TBS) • 1 or 4 theta epochs @ 0.1 Hz• Each epoch, 10 trains of 4 100Hz pulses

– Long-term depression induced by low frequency stimulation (1Hz, 15 min)

• Experimenters also blind to treatment of animals

• Fiber volley differences measured– Indirect measure of the # of axons firing

an action potential

Methods (Drug Treatments)- Fluoxetine administration

- Chronic / Subchronic treatments- 3 weeks of 18 mg/kg/day- 4 days of 18 mg/kg/day

- Dosages based on weight of mouse & average amount of water consumed/day

- Amounts consumed per mouse measured during treatments- Mice house in T7 cycle returned for 2 weeks after treatment

before testing- Desipramine administration

- 16 mg/kg 24-hours and 1-hour prior to novel object recognition paradigm

Figure 1. Aberrant light increases depression-like behavior and corticosterone levels

Figure 2. Aberrant light impairs hippocampal learning, LTP, and recognition memory

Figure 3. Chronic antidepressant administration rescues learning

Figure 4. ipRGCs mediate impairment of mood and learning by aberrant light

Conclusions

• Aberrant light can directly influence mood and learning despite normal sleep/circadian rhythms

• Light detection in ipRGCs may be mediating the deficits observed

• Antidepressant medications can alleviate the deficits caused by disruptive light environments

• T7 light cycle can serve as an alternative experimental paradigm for studying depression

Future research?• What subtypes of ipRGCs are responsible for relaying the aberrant light

information?• To what areas of the brain is the input about aberrant light being relayed;

how is it influencing those areas?• Can varying light intensities mediate different degrees of the cognitive

deficiency?• How long does aberrant light have to persist to cause deficiencies?• Why are depressive behaviors and learning influenced, but not anxiety-

related behaviors?• Which of the two antidepressants worked better and how are they

alleviating the deficiencies? • Can a change from T7 back to T24 reverse deficits as well?• How clinically relevant are these findings?

– In what situation would a person find themselves in a light pattern similar to the T7 light cycle (3.5hr light/3.5hr dark)?

Learning Objectives• How are intrinsically photosensitive retinal ganglion

cells (ipRGCs) “intrinsically photosensitive”?• What properties of ipRGCs make them suited for

relaying irradiance information over long periods of time?

• In what ways are the various subtypes of ipRGCs differentiated?

• What evidence is there to extend the functional role of some ipRGC subtypes beyond “non-image forming”?

• Despite normal circadian rhythms, what effects can aberrant light have on individuals mood and learning via ipRGCs?