The Regulatory Role Of CRH In Itch Sensation

Everyone in his lifetime has experienced acute or chronic itch.Acute itch is a protective mechanism for humankind to evade injury or danger and detect diseases earlier,while chronic itch loses its protective role which always results in insomnia,negative emotions,lose of interest,anhedonia,and eventually leads to anxiety,depression and even suicide.On the other hand,long-term mental pressure or psychiatric diseases such as anxiety and depression will also induce or aggravate pruritus dermatosis,such as atopic dermatitis,neurodermatitis,psoriasis and leukodermia.In a word,chronic itch and negative emotions could form an“itch-depression-itch”vicious cycle.So far,effective treatment for chronic itch and negative emotions is still lacking^[1].The key to this problem is to find a molecular target which could simultaneously regulate itch and negative emotions.As is known to us all,the hypothalamic-pituitary-adrenal axis?HPA?participates in individual stress reactions.Upon stressful stimuli,the hypothalamic paraventricular nucleus will release corticotropin-releasing hormone?CRH?to react to stress.Previous studies have suggested that CRH and its receptors play an important role in the“pain-depression-pain”vicious cycle^[2-4].Then,how could we establish the interactive aggravation experimental model of chronic itch and depression?Whether the decreased sensation of itch and pain in the condition of acute stress is associated with the surge in blood CRH content?Whether CRH participates in the regulation of itch?To solve these issues,we firstly established the interactive aggravation model of chronic itch and depression,and then we investigated the role of CRH and its receptors in itch regulation via pharmacological method,ELISA,immunostaining,Western-blot and in situ hybridization?ISH?.Part ? Behavioral study of the interactive aggravation model of chronic itch and depressionObjectiveTo establish the interactive aggravation model of chronic itch and depression and observe the phenomenon of the interactive aggravation of chronic itch and depression.Methods?1?The establishment of chronic itch model.Male mice were used to establish chronic itch model via neck skin administration of 0.15%DNFB for consecutive 5 days.Scratching number was counted for 30 min.Forced swimming,tail suspension and splashing experiments were conducted to examine depression behaviors while rostarod experiment was conducted to examine locomotion ability.?2?The establishment of chronic itch model.Male mice were used to establish depression model via chronic unpredicted mild stimulation?CUMS?method for consecutive 28 days.Scratching and depression behaviors were examined 28 days later.?3?The establishment of combining chronic itch and depression model.Mice were classified into three groups:A group received combining CUMS and DNFB treatment;B group received only CUMS treatment;C group received only DNFB treatment.All treatments sustained for 14 days and then depression and scratching behaviors were examined and analysed.?4?The effects of depression on acute itch.?1?CUMS mice and control mice received intradermal injection of histamine or cloroquina on the neck skin to explore the effect of chronic stress on acute itch.?2?Mice were classified into three groups:A group received combining CUMS and ketamine treatment;B group received only ketamine treatment;C group received CUMS and saline treatment.Ketamine was administrated on day 1 and 14.Depression and scratching behaviors were examined 28 days later.Results?1?Compared to control,chronic itch mice exhibited increased scratching number,increased immobility time in tail suspension and forced swimming experiment and increased grooming time in splashing experiment?P<0.01?.?2?Compared to control,CUMS model mice exhibited increased scratching number,increased immobility time in tail suspension and forced swimming experiment and increased grooming time in splashing experiment?P<0.01 or 0.05?.?3?Compared to Group B or C,CUMS and DNFB combining model mice exhibited severer depression behavior intail suspension,forced swimming and splashing experiments?P<0.01 or 0.05?,as well as more scratching number?P<0.01?.?4?Compared to control group,CUMS model mice exhibited more scratching number.Compared to another two groups,CUMS+K group exhibited less immobility time in forced swimming test and less scratching number?P<0.01?.The sole ketamine administration decreased the immobility time in forced swimming test without any effects on histamine induced scratching number.ConclusionsThe interactive aggravation model of chronic itch and depression was established and the phenomenon of the interactive aggravation of chronic itch and depression was observed.Ketamine exerts anti-depression effects and eliminated depression induced increased itch sensitivity,which will lay the foundation for subsequent studies.Part ? The regulatory role of CRH in itch sensationObjective To observe the regulatory role of CRH in itch sensation and explore its receptor mechanisms.Methods?1?ELISA and pharmacological methods were conducted to observe the effects of forced swimming on acute pain or itch and the relationship between blood CRH content and itch or pain behavior.?2?The effects of CRH on acute or chronic itch were observed via a series of administration method,such as intracerebroventricular,intrathecal,intraperitoneal and intracutaneous injection.?3?Intracerebroventricular and intrathecal administration of CRH receptor agonist or antagonist were conducted to examine the receptor mechanisms of the regulatory role of CRH in itch sensation.?4?The expression of CRH and its receptors were examined via immunostaining,ISH and western-blot methods.Results?1?ELISA tests showed that blood CRH content reached to its peak 5-10 min after forced swimming,during which time mice exhibited obvious scratching behavior and increased mechanical pain threshold.?2?Intracerebroventricular,intrathecal and intraperitoneal injection of CRH significantly inhibited acute itch;intrathecal and intraperitoneal injection of CRH significantly inhibited chronic itch.Intracutaneous injection of high dose CRH could induce scratching behavior and facilitate histamine-induced itch.?3?Intrathecal and intraperitoneal injection of CRH receptor nonselective antagonist CRF9-41 could over-ride the inhibitory effect of CRH on histamine-induced itch,suggesting that the inhibitory effect of CRH on histamine-induced itch is mediated by CRH receptor.?4?Intrathecal injection of CRH2 receptor?CRH2R?antagonist A-2B could block the inhibitory effect of CRH and enhance histamine-induced tch;intrathecal injection of CRH1 R antagonist NBI27914 exerted no effect on histamine-induced itch,suggesting that CRH inhibits itch sensation via CRH2 R while CRH1 R may facilitate itch sensation.?5?Intracerebroventricular injection of high-dose?0.2 mol/L?CRH1R agonist stresion-1 induced freezing behavior in mice while low-dose stresion-1 enhanced histamine induced itch,suggesting that low-dose CRH enhances itch sensitivity.?6?Western-blot data showed that chronic itch mice exhibited increased expression of CRH and its receptor within the spinal dorsal horn compared to the control;immunostaining data showed that chronic itch mice exhibited increased expression of CRH within the spinal dorsal horn compared to the control;ISH data showed that chronic itch mice exhibited increased expression of CRH1 R within the spinal dorsal horn compared to the control.ConclusionsCRH and its receptors?CRH1R and CRH2R?participate in the regulation of acute and chronic itch.5-10 min after acute stress,hypothalamus releases CRH to inhibit acute itch and pain.CRH2 R may mediate the inhibitory effects of CRH on acute or chronic itch while CRH1 R may mediate the facilitatory effects of CRH on itch.