These results indicate that constitutive expression of intracelluar IFN-a does not have obvious negative effects on the early-stage development of transgenic SCNT calves. IFN-a can influence the proliferation, differentiation, and function of various types of cells in the immune system and thus influence lympho-hematopoiesis. However, it has been proposed that this effect only takes place under conditions of trauma or inflammation. These factors could explain why expression of hIFN-a did not have obvious adverse effects on transgenic calves in our study. This is supported by a report in which transgenic mice expressing IFN-b displayed similar behavior, external anatomy, life span, and female fertility as wild type mice, although male transgenic mice displayed reduced fertility. The idiopathic infertilities may be caused by IFNs in the extracellular fluid, which affect the interplay between germ cells and Sertoli cells. It has also been shown that the level of IFN-a in the seminal plasma may be related to sperm production. Intracellular expression of hIFNa did not result in a high level of IFN-a in the seminal plasma in our study. However, the fertility and the viral resistance of the transgenic calves need to be further determined. The calf is now 17 months old and continued GFP expression has been shown by immunofluorescence in an ear biopsy and in fibroblasts derived from the animal. In conclusion, we constructed a vector with a hIFN-a gene cassette. Stably transfected bovine fetal fibroblasts were obtained, and hIFN-a transgenic embryos were produced by SCNT. Two male cloned calves were born. Expression of intracellular hIFN-a conferred viral resistance on transfected bovine fetal fibroblasts and did not significantly affect the full development of SCNT embryos. These data suggest that IFN-a transgenic technology may provide a revolutionary way to achieve disease-control measures through elite breeding of livestock. The cholinergic neurons of the medial septal nucleus send their axons to the hippocampus providing it with a robust modulatory input mediated by their neurotransmitter, acetylcholine. Although not numerous, their significance includes critical functions in processes of learning, memory, attention, and sleep. Degeneration and/ or malfunction of the septohippocampal cholinergic system is thought to contribute to cognitive decline of old age and to memory loss characteristic of Alzheimer’s disease. Lesions of this pathway in laboratory animals impair learning and memory in various tests. Because these abnormalities are related to the compromised cholinergic neurotransmission and since the septohippocampal pathway is the main extrinsic source of ACh to the hippocampus, multiple experimental strategies aimed at restoring the content of this neurotransmitter in the hippocampus have been tried. For example, grafts of ACh-producing cells ameliorated some lesion-evoked cognitive deficits. In an effort to correct similar deficits in AD, a number of therapeutic approaches have also been undertaken to increase the levels of brain ACh in humans. Some of these strategies are based on the use of acetylcholinesterase inhibitors, to reduce the rate of degradation of ACh in brain. Other approaches include the use of growth factors known to be neurotrophic for cholinergic neurons and to be effective in animal models – NGF being the best characterized -and clinical trials on NGF gene therapy in an attempt to increase the functional state and survival of cholinergic neurons in AD patients have been initiated. We have previously shown that bone morphogenetic protein 9 is an inducer of the cholinergic phenotype of septal neurons in culture and in embryonic brain in vivo and a maintenance factor for these cells. Moreover, in fetal septal cell cultures, BMP9 induces the expression of multiple genes, a large fraction of which belong to the transcriptome of the septal cholinergic neurons, indicating that BMP signaling may participate in the maturation of these cells more broadly, i.e. beyond their neurotransmitter specification. Interestingly, several BMP9-induced genes encode proteins with trophic activities for cholinergic neurons, suggesting that BMP signaling helps to generate a favorable milieu for these cells. Indeed, we have recently demonstrated that some of the effects of BMP9 are apparently mediated by autocrine action of NGF that is induced in these neurons in response to BMP9. Multiple BMPs and their antagonists also function in controlling hippocampal development and modulate adult neurogenesis that is characteristic to this brain region. In the present study we found that intracerebroventricular infusion of BMP9 prevents the loss of cholinergic neurons, as defined by their expression of the ACh-synthesizing enzyme choline acetyltransferase, in the septum following unilateral fimbria-fornix transection in mice and upregulates the expression of hippocampal NGF and its receptors NGFR-p75 and TRKA. To assess the in vivo activity of BMP9 on injured adult septal cholinergic neurons, we performed unilateral fimbria-fornix transections in mice and treated them with varying doses of BMP9 administered via continuous ICV infusion over a 6-day period. Immunohistochemical staining of the septum with an antibody for CHAT was then used to determine the effects of the lesion and of BMP9. Serial images of the septal/diagonal band area from CHAT-immunostained histological sections were processed for cell number quantification within the region of interest encompassing the individual left and right medial septal nuclei defined by triangular shapes shown in Figure 1 as Mecamylamine hydrochloride described in the methods.