Distribution of heat shock protein 108 mRNA in the chicken central nervous system Dong Hoon Shin a , Hyun Joon Kim b , Hwa Young Lee a , Kyung Hoon Lee c , Gye Sun Jeon d , Je Hoon Seo d , Sang Ho Baik d , Sa Sun Cho d, e, * a Department of Anatomy, Dankook University College of Medicine, Chonan, South Korea b Department of Anatomy, Gyeongsang National University College of Medicine, Chinju, South Korea c Department of Anatomy, Sungkyunkwan University College of Medicine, Suwon, South Korea d Department of Anatomy, Seoul National University, College of Medicine, Yongon-Dong 28, Seoul 110-799, SouthKorea e Clinical Research Institute, Seoul National University Hospital, Seoul, South Korea Received 10 January 2000; received in revised form 14 February 2000; accepted 16 February 2000 Abstract The constitutive expression of heat shock protein 108 (HSP108) mRNA is mapped in a normal chicken central nervous system using in situ hybridization technique. HSP108 mRNAs were found to be mainly localized in the small neuroglial cells of various regions of the brain, although some neuronal cells also showed positive signals. This tendency is observed to be more marked in the cerebellum; HSP108 signals were not found in the Purkinje cells, but in Bergmann glial cells and oligodendrocytes. Although neuronal cells in the deep cerebellar nuclei and the molecular layer showed occasional HSP108 signals, the expression pattern of HSP108 mRNA is different from homologous HSP90 that is mostly expressed in neurons, but rather similar to that of TfBP immunoreactivity, a new member of the HSP108 family. The constitutive neuroglial localization of HSP108 could suggest that HSP108 may play an important role in the normal metabolism of neuroglial cells in the chicken brain. q 2000 Elsevier Science Ireland Ltd. All rights reserved. Keywords: In situ hybridization; Brain; Heat shock protein; Chicken; Bergmann glial cell; Oligodendrocyte Heat shock protein (HSP) is known to include a series of stress proteins induced by several different stressful condi- tions such as heat, ischemia, in¯ammation, oxidative stress, and even behavioral and psychological stresses [12,13]. However, the results of several studies suggest that they are not restricted to expression in stress conditions, but rather are also expressed under normal conditions [4,5,7,19]. HSP108, the avian homologue of the mammalian GRP94 family of stress-regulated proteins [6,14], was originally puri®ed from chicken oviduct [11]. Although transferrin binding protein (TfBP), which also belongs to the HSP108 family was extensively studied in our labora- tory [1], comparatively little work has been carried out on the expression of HSP108 mRNA in the chicken brain. Thus in the present study, we ®rst utilized riboprobes speci®c to HSP108 mRNA to reveal the constitutional distribution of HSP108 mRNA in the chicken central nervous system (CNS). The brains of adult white leghorn chicken were used in this study. After the tissues were sliced into 12 mm sections on a cryostat, the sections were ®xed in 4% (w/v) paraformalde- hyde solution and treated with chloroform for the purpose of protein removal. The animals used in this experiment were treated in accordance with the `Principles of Laboratory Animal Care' (NIH publication No. 86-23, revised 1985). Total cellular RNA was extracted from white leghorn chick- ens aged embryonic day 18 (E18) using the guanidine thio- cyanate (GTC) method [17]. HSP108 cDNA was ampli®ed by reverse-transcription polymerase chain reaction (RT- PCR) method using the oligonucleotide primers such as 5 0 HSP108 primer: CCA GTT TGG TGT TGG CTT TT and 3 0 HSP108 primer: CCT CCT TTG CTT CCT CCT CT. The design of these PCR primers was based on previous cloning results [3,11], and the predicted size of the ampli®ed chicken HSP108 cDNA was 323bp. The PCR products were cloned into T easy vector (Promega) and sequenced by the dideox- ynucleotide chain termination method applying the T7 sequencing kit (Pharmacia). Digoxigenin-11-UTP (Boehrin- ger-Mannheim) labeled antisense HSP108 cRNA probe was Neuroscience Letters 283 (2000) 181±184 0304-3940/00/$ - see front matter q 2000 Elsevier Science Ireland Ltd. All rights reserved. PII: S0304-3940(00)00967-8 www.elsevier.com/locate/neulet * Corresponding author. Tel.: 182-2-740-8204; fax: 182-2-745- 9528. E-mail address: chossn@snu.ac.kr (S.S. Cho).