The LH,the“wakefulness center”,is characterized by:orexinergic neurons,that produce orexin-A and B,and MCHneurons.Both,OX and MCHneurons,are critical regulators of wakefulness and sleep being wake-active and sleep-active,respectively.OX and MCHneurons are intermingled with GABAergic neurons maximally active during sleep and by their projections could inhibit OXneurons,to promote sleep.On the other hand,cannabinoids,the primary active agent in drugs of abuse,tend to generate a distorted sense of time.Endocannabinoids(EC)(2AGandAEA)show a circadian variation in the brain.Here we study the role of ECsystem in the modulation of wakefulness,hypothesizing a light-mediate ECsynthesis and release in the LH by means of the collateral-way of retinohypothalamic tract(RHT)described by Leak&Moore.The RHT is made by projections of pRGcells which detect light directly by using melanopsin,a photopigment responsive to blue-light.Glutamate is the primary neurotransmitter released at RHTprojections.Our previous studies suggest that ECsignaling exert a powerful modulation of OXAneurons,which synthesize 2AG and are innervated by glutamatergic/GABAergic inputs carrying CB1receptor.However lack of data exist despite to the expression of CB1 on RHTprojections to LH and on its activation following a possible light-mediated shift of EClevels.In order to unravel the question we identified RHTprojections to LH by intravitreal injection of anterograde tracerCTB performed in adult mice maintained at 12D/L.At 7 days after injection the CTBtracing were revealed by immunohistochemistry coupled to OXA,MCH,GAD65,CB1 revelation.We observed the majority of glutammatergicCB1/CTB and some gabaergicCB1/GAD65projections around OXAneurons as confirmed by CB1/OXA and CB1/DAGLα-immunoelectronmicroscopy. Moreover,a functional light-mediated activation of OXAneurons was tested by blue light-pulse excitation of retina at CT14.After 90min of dark,the LH was processed for c-Fos-revelation coupled to OXA,MCH,GAD65immunostaining.A strong c-Fos-expression was observed in OXAneurons.We supposed that synthesis and release of 2AG could be induced by glutamatergic light-mediated-activation of OXAneurons.2AG could,in turn,induce self-inhibition of OXAneurons as effect of CB1activation on RHTglutammatergic afferences.This hypothesis is supported by significant increase of 2AGlevel in LH up to 15min of light-pulse and concurrently activation of Ca2+indicator(Fluo-4-dextran)presinaptically to OXneurons.
Retinohypothalamic projections to LH are involved in the light-mediated modulation of wakefulness through interaction with orexinergic and endocannabinoid systems / R., Imperatore; A., Iovine; A., DI NUNZIO; Ferrandino, Ida; V., DI MARZO; L., Cristino. - (2011), pp. C138-C138. (Intervento presentato al convegno 8th IBRO World Congress of Neuroscience tenutosi a Firenze nel 14-18 Luglio 2011).
Retinohypothalamic projections to LH are involved in the light-mediated modulation of wakefulness through interaction with orexinergic and endocannabinoid systems
FERRANDINO, IDA;
2011
Abstract
The LH,the“wakefulness center”,is characterized by:orexinergic neurons,that produce orexin-A and B,and MCHneurons.Both,OX and MCHneurons,are critical regulators of wakefulness and sleep being wake-active and sleep-active,respectively.OX and MCHneurons are intermingled with GABAergic neurons maximally active during sleep and by their projections could inhibit OXneurons,to promote sleep.On the other hand,cannabinoids,the primary active agent in drugs of abuse,tend to generate a distorted sense of time.Endocannabinoids(EC)(2AGandAEA)show a circadian variation in the brain.Here we study the role of ECsystem in the modulation of wakefulness,hypothesizing a light-mediate ECsynthesis and release in the LH by means of the collateral-way of retinohypothalamic tract(RHT)described by Leak&Moore.The RHT is made by projections of pRGcells which detect light directly by using melanopsin,a photopigment responsive to blue-light.Glutamate is the primary neurotransmitter released at RHTprojections.Our previous studies suggest that ECsignaling exert a powerful modulation of OXAneurons,which synthesize 2AG and are innervated by glutamatergic/GABAergic inputs carrying CB1receptor.However lack of data exist despite to the expression of CB1 on RHTprojections to LH and on its activation following a possible light-mediated shift of EClevels.In order to unravel the question we identified RHTprojections to LH by intravitreal injection of anterograde tracerCTB performed in adult mice maintained at 12D/L.At 7 days after injection the CTBtracing were revealed by immunohistochemistry coupled to OXA,MCH,GAD65,CB1 revelation.We observed the majority of glutammatergicCB1/CTB and some gabaergicCB1/GAD65projections around OXAneurons as confirmed by CB1/OXA and CB1/DAGLα-immunoelectronmicroscopy. Moreover,a functional light-mediated activation of OXAneurons was tested by blue light-pulse excitation of retina at CT14.After 90min of dark,the LH was processed for c-Fos-revelation coupled to OXA,MCH,GAD65immunostaining.A strong c-Fos-expression was observed in OXAneurons.We supposed that synthesis and release of 2AG could be induced by glutamatergic light-mediated-activation of OXAneurons.2AG could,in turn,induce self-inhibition of OXAneurons as effect of CB1activation on RHTglutammatergic afferences.This hypothesis is supported by significant increase of 2AGlevel in LH up to 15min of light-pulse and concurrently activation of Ca2+indicator(Fluo-4-dextran)presinaptically to OXneurons.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
