title: Selective lesioning of nucleus incertus with corticotropin releasing factor-saporin conjugate.
abstract: The nucleus incertus (NI), a brainstem nucleus found in the pontine periventricular grey, is the primary source of the neuropeptide relaxin-3 in the mammalian brain. The NI neurons have also been previously reported to express several receptors and neurotransmitters, including corticotropin releasing hormone receptor 1 (CRF₁) and gamma-aminobutyric acid (GABA). The NI projects widely to putative neural correlates of stress, anxiety, depression, feeding behaviour, arousal and cognition leading to speculation that it might be involved in several neuropsychiatric conditions. On the premise that relaxin-3 expressing neurons in the NI predominantly co-express CRF₁ receptors, a novel method for selective ablation of the rat brain NI neurons using corticotropin releasing factor (CRF)-saporin conjugate is described. In addition to a behavioural deficit in the fear conditioning paradigm, reverse transcriptase polymerase chain reaction (RT-PCR), western blotting (WB) and immunofluorescence labelling (IF) techniques were used to confirm the NI lesion. We observed a selective and significant loss of CRF₁ expressing cells, together with a consistent decrease in relaxin-3 and GAD65 expression. The significant ablation of relaxin-3 positive neurons of the NI achieved by this lesioning approach is a promising model to explore the neuropsychopharmacological implications of NI/relaxin-3 in behavioural neuroscience.
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University Health System, 117597, Singapore; Neurobiology and Ageing Programme, Life Sciences Institute, National University of Singapore, 117456, Singapore; Singapore Institute for Neurotechnology (SINAPSE), 117456, Singapore.
authors.forename: Liying Corinne
authors.initials: LC
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University Health System, 117597, Singapore; Neurobiology and Ageing Programme, Life Sciences Institute, National University of Singapore, 117456, Singapore; Singapore Institute for Neurotechnology (SINAPSE), 117456, Singapore.
authors.forename: Ramamoorthy
authors.initials: R
authors.lastname: Rajkumar
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University Health System, 117597, Singapore; Neurobiology and Ageing Programme, Life Sciences Institute, National University of Singapore, 117456, Singapore; Singapore Institute for Neurotechnology (SINAPSE), 117456, Singapore. Electronic address: gavin_dawe@nuhs.edu.sg.
authors.forename: Gavin Stewart
authors.initials: GS
authors.lastname: Dawe
mesh_terms: D000818:Animals; D003213:Conditioning, Psychological / Q000187:drug effects; D003346:Corticotropin-Releasing Hormone / Q000378:metabolism / Q000633:toxicity*; D004597:Electroshock / Q000009:adverse effects; D005239:Fear / Q000187:drug effects; D005786:Gene Expression Regulation / Q000502:physiology; D005904:Glial Fibrillary Acidic Protein / Q000235:genetics / Q000378:metabolism; D000922:Immunotoxins / Q000633:toxicity*; D008297:Male; D011149:Pons / Q000293:injuries* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D018019:Receptors, Corticotropin-Releasing Hormone / Q000235:genetics / Q000378:metabolism; D012065:Relaxin / Q000235:genetics / Q000378:metabolism*; D054789:Ribosome Inactivating Proteins, Type 1 / Q000633:toxicity*; D000076985:Saporins; D013997:Time Factors; D014365:Tryptophan Hydroxylase / Q000235:genetics / Q000378:metabolism
chemical_list: D005904:Glial Fibrillary Acidic Protein; D000922:Immunotoxins; D018019:Receptors, Corticotropin-Releasing Hormone; D054789:Ribosome Inactivating Proteins, Type 1; D012065:Relaxin; D003346:Corticotropin-Releasing Hormone; D014365:Tryptophan Hydroxylase; C504569:tph2 protein, rat; D000076985:Saporins
keywords: CRF–saporin; Fear conditioning; Lesion; Nucleus incertus; Rat; Relaxin-3