Lee S, Smith G W, Vale W, Lee K F, Rivier C
Clayton Foundation Laboratories for Peptide Biology, The Salk Institute, La Jolla, California 92037, USA.
Alcohol Clin Exp Res. 2001 Mar;25(3):427-33.
The purpose of this work was to determine the influence of acute alcohol treatment, injected intraperitoneally, on the hypothalamic-pituitary-adrenal axis of mice that lack type 1 receptor for corticotropin-releasing factor (CRFR1).
CRFR1-deficient (CRFR1-/-), heterozygous (CRFR1+/-), and wild-type (CRFR1+/+) mice were generated and maintained under standard conditions. Homozygous, heterozygous, and wild-type offspring were identified by polymerase chain reaction analysis of tail DNA. Experiments were performed on 9- to 16-week-old male and female mice. All blood samples were obtained by rapid decapitation of conscious mice conducted between 10 AM-12 PM. Blood sample collection was completed within 20 to 30 sec of disturbing the animals, and all samples were terminal. Preliminary experiments were conducted to determine the time-course of the ACTH and hypothalamic responses to alcohol in all three groups of mice, and a single time point (30 min and 2 hr, respectively), corresponding to peak responses, was chosen to measure the corresponding parameters in all subsequent studies.
In vehicle-injected animals, basal ACTH and corticosterone levels were statistically comparable in heterozygotes and mice with a null allele for the CRFR1 gene, although values of this latter hormone were slightly lower in the mutants. Alcohol (4.0 g/kg) elicited the expected significant (p < 0.01) increase in plasma ACTH and corticosterone levels in heterozygous mice. These responses were virtually abolished or markedly decreased, respectively, in CRFR1-deficient animals. As previously reported, constitutive CRF mRNA levels were elevated in the paraventricular nucleus (PVN) of the hypothalamus in mice that lacked CRFR1, compared to wild-type control mice. Interestingly, this was not the case for transcripts of the immediate early gene NGFI-B. When measured 2 hr after alcohol, PVN NGFI-B gene expression was significantly (p < 0.01) increased in both control and mutant mice, as were CRF mRNA levels in mutant mice, but the hypothalamic responses of the mutants were larger (p < 0.01) than those of the control mice. This difference may be due, at least in part, to the lack of steroid feedback in the mutants.
These results indicate that although the intraperitoneal injection of alcohol remains capable of eliciting PVN CRF neuronal activation in mice that lack CRFR1, the ACTH and corticosterone responses are significantly blunted, a phenomenon believed to be due to the lack of CRFR1 in the pituitary of these animals.
本研究旨在确定腹腔注射急性酒精对缺乏促肾上腺皮质激素释放因子1型受体（CRFR1）的小鼠下丘脑-垂体-肾上腺轴的影响。
培育CRFR1基因缺陷型（CRFR1-/-）、杂合子型（CRFR1+/-）和野生型（CRFR1+/+）小鼠，并在标准条件下饲养。通过对尾DNA进行聚合酶链反应分析来鉴定纯合子、杂合子和野生型后代。对9至16周龄的雄性和雌性小鼠进行实验。所有血样均通过在上午10点至12点之间对清醒小鼠进行快速断头采集。在干扰动物后20至30秒内完成血样采集，所有样本均为终末样本。进行初步实验以确定三组小鼠中促肾上腺皮质激素（ACTH）和下丘脑对酒精反应的时间进程，并选择一个对应峰值反应的单一时间点（分别为30分钟和2小时）来测量所有后续研究中的相应参数。
在注射溶剂的动物中，杂合子小鼠和CRFR1基因无功能等位基因的小鼠的基础ACTH和皮质酮水平在统计学上具有可比性，尽管后一种激素在突变体中的值略低。酒精（4.0 g/kg）使杂合子小鼠的血浆ACTH和皮质酮水平出现预期的显著升高（p < 0.01）。在CRFR1基因缺陷型动物中，这些反应分别几乎完全消失或明显减弱。如先前报道，与野生型对照小鼠相比，缺乏CRFR1的小鼠下丘脑室旁核（PVN）中组成型促肾上腺皮质激素释放因子（CRF）mRNA水平升高。有趣的是，立即早期基因NGFI-B的转录本情况并非如此。在酒精注射2小时后测量时，对照小鼠和突变体小鼠的PVN中NGFI-B基因表达均显著增加（p < 0.01），突变体小鼠中的CRF mRNA水平也显著增加，但突变体小鼠的下丘脑反应比对照小鼠更大（p < 0.01）。这种差异可能至少部分归因于突变体中缺乏类固醇反馈。
这些结果表明，尽管腹腔注射酒精仍能够在缺乏CRFR1的小鼠中引发PVN中CRF神经元的激活，但ACTH和皮质酮反应明显减弱，这种现象被认为是由于这些动物垂体中缺乏CRFR1所致。
