On the role of forebrain cholinergic innervation for phencyclidine-induced behaviors and gene expression patterns

摘要

The basalo-cortical cholinergic innervation reaches neuraltargets in cortex cerebri where it modulatesthe incoming sensory information, regulating arousal, attention, vigilance, memory and cognition.Progressive degeneration of the cholinergic neurons in the basal forebrain is considered a hallmark ofAlzheimer’s disease related to cognitive impairment in this condition. Dysregulation of cholinergicneurotransmission has also been implicated in schizophrenia, a chronic and debilitatingneurodevelopmental disorder characterized by positive and negative symptomsand cognitiveimpairments, with deficits in thought processes, perceptions and emotional responsiveness. Modelingspecific disruption of cholinergic function allows studies of its behavioral and molecular consequencesand may contribute to the developmentof future therapies for Alzheimer’s disease and Schizophrenia.To investigate the role of the cholinergic system, we carried out uni-and bilateral cholinergicdenervation of cortex cerebri in male Lister hooded rats using the selective immunotoxin 192 IgG-saporin to specifically target the cholinergic neurons of nucleus basalis of magnocellularis (NBM). IgG-saporin effectively removed cholinergic afferents to the cortical mantle. Intact and denervated rats werechallenged with phencyclidine (PCP), a non-competitive NMDA receptor antagonist, which is known toproduce schizophrenia-like psychosis in humans and hence used to model aspects of schizophrenia inrats. Experiments were conducted to determine the effects on behavior and to shed light on theunderlying molecular mechanisms.Negative symptoms of schizophrenia include social withdrawal, which is studied in animal models usingsocial interaction tests. We found that cortical cholinergic denervation lead to a significant reduction inthe duration of active social interaction in pairs of lesioned animals versus pairs of sham-operatedcontrols. After an acute dose low of PCP (1 mg/kg, s.c.), there was a marked decrease in active socialinteraction for both groups, such that there was no longer a difference between lesioned and controlanimals. To evaluate cognitive impairments, particularly those reflecting declarative memory, a novelobject recognition test was used. Neither cholinergic denervation alone, nor an acute PCP dose (1 mg/kg,s.c.) alone, blocked the ability of rats to recognize a novel object. However, animals lacking corticalcholinergic innervation and challenged with PCP were no longer able to recognize the novel object(Paper I).Behavioral paradigms have typically been analyzed manually, though automated hardware andsoftware systems have been introduced to speed up the process and eliminate subjective errors. How-ever, the reliability and accuracy of computerized scoring needs to be verified for each specificparadigm. For novel object recognition, the program scoring reduced analysis time for the task, butneeded manual corrections to avoid erroneous data points. Manual for scoring of social interactionrevealed that this labor intensive approach was more nuanced and allowed to discern complex behaviors,whereas the automated scoring system registered more global interaction patterns. It is concluded thatmanual and computerized techniques for scoring social interaction offercomplimentary information onvarious aspects of this complex behavioral task (Paper II).The next studies were aimed to examine the bases of the behavioral responses to cortical cholinergicdenervation and PCP challenge at the molecular level. PCP-induced neuronal activation was mappedusing quantitativein situhybridization of the neuronal immediate early genec-FosmRNA levels.Transcription ofc-Fosresponds to many different forms of activation, including stress and toxins. Wefound that two doses of PCP used (2 and 3 mg/kg, s.c.) caused a marked increase in neuronalc-FosmRNA expression at 30 and 60 min after PCP administration, though these doses did not alter the levelsof BDNF or Nogo receptor mRNA. Importantly, PCP responses were markedly dampened incholinergically denervated regions of cortex cerebri (Paper III).RNA-Sequencing was then employed to understand gene regulatory processes at a global level insomatosensory cortex with regard to effects of chronic denervation and/or a PCP challenge (1 and 3mg/kg, s.c.). A first round of analysis focused on the genes most significantly altered by denervation(Egr1, Dusp6, Ier2, Nr4a1), PCP treatment (Cyr61, Bcl6b, Apold1, Dusp1) and those altered over time(Sox9, Coq10b, Zfp189, Rnf39) (Paper IV).The most common response was for mRNA levels toincrease in response to PCP and for cholinergic denervation to dampen these increases.The findings presented in this thesis support a role for the cholinergic system in assigning significance toincomingstimuli as exemplified by regulating the response of cortical neurons to a PCP challenge. Thefindings are also compatible with a proposed involvement of cholinergic dysfunction in schizophrenia.