To study the mechanisms that activate expression of the atrial natriuretic factor (ANF) gene during pressure-induced hypertrophy, we have developed and characterized an in vivo murine model of myocardial cell hypertrophy. We employed microsurgical techniques to produce a stable 35-to 45-mmHg pressure gradient across the thoracic aorta of the mouse that is associated with rapid and transient expression of an immediate-early gene program (c-fos/cjun/junB/Egr-1/nur-77), an increase in heart weight/body weight ratio, and up-regulation of the endogenous ANF gene. These responses that are identical to those in cultured cell and other in vivo models of hypertrophy. To determine whether tissue-specific and inducible expression of the ANF gene can be segregated, we used a transgenic mouse line in which 500 base pairs of the human ANF promoter region directs atrial-specific expression of the simian virus 40 large tumor antigen (T antigen), with no detectable expression in the ventricles. Thoracic aortic banding of these mice led to a 20-fold increase in the endogenous ANF mRNA in the ventricle but no detectable expression of the T-antigen marker gene. This result provides evidence that atrial-specific and inducible expression of the ANF gene can be segregated, suggesting that a distinct set of regulatory cis sequences may mediate the up-regulation of the ANF gene during in vivo pressure overload hypertrophy. This murine model demonstrates the utility of microsurgical techniques to study in vivo cardiac physiology in transgenic mice and should allow the application of genetic approaches to identify the mechanisms that activate ventricular expression of the ANF gene during in vivo hypertrophy.In response to diverse stimuli, such as hypertension, valvular heart disease, and endocrine disorders, the myocardium adapts to increased workloads through the hypertrophy of individual muscle cells (for a review, see refs. 1 and 2). Although the signaling mechanisms that mediate the hypertrophic response of cardiac muscle cells remain unclear, transcriptional activation of cardiac target genes, including contractile proteins and embryonic markers, appears to play a pivotal role in this adaptive response (3, 4). In this regard, the reactivation of atrial natriuretic factor (ANF) gene expression in ventricular cells occurs in response to diverse hypertrophic stimuli (genetic, hormonal, volume overload, pressure overload, hypertension, etc.) in multiple species (5-11), including humans, and could be considered one of the conserved features of ventricular cell hypertrophy.To study the transcriptional regulation of cardiac genes, workers in our laboratory (3, 12, 13) and others (14-16) have extensively characterized cultured myocardial cell models in which several features of hypertrophy can be induced after stimulation with defined agents, such as a-adrenergic agonists (3, 12, 14-17) or endothelin 1 (13). In this model, the inducibility of a constitutively expressed contractile protein gene, myosin light chain 2 (MLC-2), is mediat...

Chronic regulation of the cardiovascular system by atrial natriuretic factor was investigated by generating transgenic mice with elevated hormone levels in the systemic circulation. A fusion gene comprising the mouse transthyretin promoter and mouse atrial natriuretic factor structural sequences was designed so as to target hormone expression to the liver. Hepatic expression of atrial natriuretic factor was detectable as early as embryonic day 15 in transgenic animals. In adult transgenic mice, plasma immunoreactive atrial natriuretic factor concentration was elevated at least eightfold as compared with nontransgenic littermates. The mean arterial pressure of conscious transgenic mice was 75.5 ±0.9 mm Hg, significantly less than that of nontransgenic siblings (103.9±2.0 mm Hg). This difference in mean arterial pressure was not accompanied by significant changes in several other physiological parameters, including heart rate, plasma and urinary electrolytes, water intake, and urine volume. This study demonstrates that a chronic elevation of plasma atrial natriuretic factor decreases arterial blood pressure without inducing diuresis and natriuresis in transgenic mice and also illustrates the value of the transgenic approach for the study of the cardiovascular system. (Hypertension 1990;16:301-307)

Nerve growth factor (NGF) supports the survival of developing sympathetic and a subpopulation of sensory neurons. In the adult it participates in maintenance of the neurotransmitter phenotype of responsive neurons. The amount of NGF synthesized by a given target tissue determines its final innervation density; those developing neurons that fail to receive sufficient NGF undergo apoptosis. In order to examine the ramifications of this principle in the context of a specific target organ, a transgenic mouse model was developed in which NGF expression was increased in developing and adult cardiac tissue by placing a NGF minigene under the transcriptional control of the cardiac-specific alpha-myosin heavy chain promoter. Transgenic mice developed cardiac enlargement secondary to both an increase in myocardial mass and the presence of an abundant ectopic cell population. Immunohistochemical analyses with the neural marker S-100 revealed staining of a subpopulation of ectopic cells, suggesting their derivation from the neural crest. Whereas immunostaining for the neuronal-specific protein neuron-specific enolase demonstrated labeling of another subpopulation of ectopic cells within the heart. Measurements of cardiac tissue catecholamine levels revealed a marked elevation in transgenic mice, consistent with sympathetic hyperinnervation. Analysis of mediastinal sympathetic ganglia revealed increases in both the size and the number of neurons. In this model, increased expression of NGF produced hyperinnervation of the heart, pathological cardiac growth, and the recruitment and/or expansion of an ectopic, neural crest-derived cell type.