Abstract-The renin-angiotensin system plays a critical role in the control of blood pressure (BP), and its hyperactivity is associated with the development and maintenance of hypertension. Although traditional pharmacological therapies targeted toward the inhibition of the renin-angiotensin system are effective in the control of this disease, they pose significant limitations. We used an antisense gene delivery strategy to circumvent these limitations and established that a single intracardiac administration of angiotensin type 1 receptor antisense (AT 1 R-AS) causes permanent prevention of hypertension in the spontaneously hypertensive rat (SHR), an animal model of primary human hypertension. Our objectives in this study were 2-fold: to determine (1) whether the targeting of angiotensin I-converting enzyme (ACE) mRNA by a similar antisense strategy would prevent the SHR from developing hypertension and (2) whether the antihypertensive phenotype is transmitted to the offspring from the antisense-treated parents. Administration of a retroviral vector containing ACE antisense (LNSV-ACE-AS) caused a modest yet significant attenuation of high BP (Ϸ15Ϯ2 mm Hg) exclusively in the SHR. This was associated with a complete prevention of cardiac and renovascular pathophysiological alterations that are characteristic of hypertension. Like their parents, the F 1 generation offspring of the LNSV-ACE-AS-treated SHR expressed lower BP, decreased cardiac hypertrophy, and normalization of renal arterial excitation-coupling compared with offspring derived from the LNSV-ACE-tS (truncated sense)-treated SHR. In addition, the endothelial dysfunction commonly observed in the SHR renal arterioles was significantly prevented in both parents and offspring of the LNSV-ACE-AS-treated SHR. Polymerase chain reaction followed by Southern analysis revealed that the ACE-AS was integrated into the SHR genome and transmitted to the offspring. These observations suggest that transmission of ACE-AS by retroviral vector may be responsible for the transference of normotensive phenotypes in the SHR offspring. Key Words: SHR Ⅲ viral delivery Ⅲ hypertension Ⅲ cardiac hypertrophy Ⅲ renovascular responsiveness H ypertension is a complex disease that manifests as chronically high blood pressure (BP). It is a major risk factor in many cardiovascular pathophysiological states, including arteriosclerosis, stroke, heart failure, coronary artery disease, and progressive renal damage. 1-3 Overwhelming evidence has established that a dysfunctional renin-angiotensin system (RAS) is one of the many physiological alterations that contribute to the development and establishment of hypertension. 4 -6 This conclusion is based on the fact that the traditional pharmacological therapies targeted to inhibit the activity of the RAS are a highly successful strategy for the treatment and management of this disease in a significant population of hypertensive patients. [7][8][9][10] Despite their success, traditional pharmacological agents, such as angiotensin I-converting en...

Immunocytochemical staining with antibodies to the class III intermediate filament protein peripherin reveals discrete subpopulations of neurons and nerve fibres throughout the rat central nervous system. Some of these fibres enter the cerebellar granular and molecular layers. Here we use light and electron microscopic immunocytochemistry and confocal fluorescence microscopy to identify the peripherin positive fibres in the molecular layer of the cerebella of various mammals. (1) The peripherin positive fibres in the molecular layer have morphological attributes of climbing fibres, and peripherin positive fibres are also detected in the olivo-cerebellar tract. Furthermore peripherin positive neurons can be seen in the inferior olive, from which climbing fibres originate. (2) The peripherin positive molecular layer fibres rapidly degenerate in rats treated with 3-acetylpyridine (3-AP), a reagent which destroys neurons in the inferior olive, and the time course of degeneration of these mirrors that previously described for 3-AP induced destruction of climbing fibres. (3) Cerebella of other mammal species tested (mouse, rabbit, pig, cow and human) revealed a similar peripherin staining pattern in the cerebellum, including fibres in the molecular layer with the morphology of climbing fibres. (4) We also noted peripherin positive spinocerebellar and vestibulocerebellar mossy fibres in the cerebellar granular layer of folia known to receive these inputs. (5) A subset of perivascular nerve fibres are also peripherin positive. These results show that peripherin is a useful marker for mammalian cerebellar climbing fibres, and that a subset of morphologically distinct cerebellar mossy fibres are also peripherin positive.

Abstract-The renin-angiotensin system plays a critical role in the control of blood pressure, and its hyperactivity is associated with the development of human primary hypertension. Because low-dose angiotensin I-converting enzyme (ACE) inhibitors cause small reductions in blood pressure that are associated with the complete reversal of altered vascular pathophysiology, our objective in this study was to determine whether ACE antisense (ACE-AS) gene delivery prevents alterations in renal vascular physiology in the parents and F 1 offspring of AS-treated spontaneously hypertensive rats (SHR). A single bolus intracardiac injection of ACE-AS (2ϫ10 8 colony-forming units) in SHR neonates caused a modest (18Ϯ3 mm Hg, nϭ7 to 9) lowering of blood pressure, which was maintained in the F 1 generation offspring (nϭ7 to 9). Alterations in renal vascular reactivity, electrophysiology, and [Ca 2ϩ ] i homeostasis are underlying mechanisms associated with the development and establishment of hypertension. Renal resistance arterioles from truncated ACE sense-treated SHR showed a significantly enhanced contractile response to KCl and phenylephrine (nϭ24 rings from 6 animals, PϽ0.01) and significantly attenuated acetylcholine-induced relaxations (nϭ24 rings from 6 animals, PϽ0.01) compared with arterioles from ACE-AS-treated SHR. In addition, compared with cells dissociated from arterioles of ACE-AS-treated SHR, cells from truncated ACE sense-treated animal vessels had a resting membrane potential that was 22Ϯ4 mV more depolarized (nϭ38, PϽ0.01), an enhanced L-type Ca 2ϩ current density Key Words: hypertension, renal Ⅲ calcium channels Ⅲ potassium channels Ⅲ arterioles Ⅲ gene therapy H ypertension is a complex disease that is manifested as chronically high blood pressure (BP) and is a major risk factor in many cardiovascular pathophysiological states, including arteriosclerosis, stroke, heart failure, coronary artery disease, and progressive renal damage. 1-3 Evidence has established that a dysfunctional renin-angiotensin system (RAS) is one of the many physiological alterations that contribute to the development and maintenance of hypertension. 4 -6 This is based on the fact that pharmacological interruption in the activity of the RAS has proven to be highly successful in the treatment and management of hypertension in a significant population of hypertensive patients. 7-10 However, this pharmacological intervention has major limitations, which include compliance, side effects, and relatively short duration of antihypertensive effects. 11,12 As a result of these limitations, we have begun to use an antisense (AS) gene therapy approach to determine whether targeting RAS at a genetic level would be a step forward in the long-term control of hypertension. These studies have revealed that a single intracardiac administration of retroviral vector containing angiotensin II type 1 receptor (AT 1 R) AS results in long-term prevention of high BP in the spontaneously hypertensive rat (SHR), which is an animal model associated with stud...