Latent inhibition and conditioning in rat strains which show differential prepulse inhibition.

Latent inhibition (LI) is the retardation of associative conditioning resulting from preexposure of the conditioned stimulus (CS) alone prior to conditioning. Schizophrenic patients show deficient prepulse inhibition (PPI) and, at least acutely, deficient LI as well. We recently found that Brown Norway (BN) rats show a PPI deficit compared to Wistar-Kyoto (WKY) rats. If PPI and LI depend on neural processes with common genetic substrates, then LI should be deficient in BN rats as well. Here, LI of a conditioned taste aversion was examined in BN and WKY rats. One group from each strain was preexposed to a saccharin-flavored solution (CS) the day prior to conditioning. For taste aversion conditioning, these two groups again consumed saccharin and were injected with lithium chloride (unconditioned stimulus) 10 min later. A second group from each strain was not preexposed to the CS and was treated identically during conditioning, while a third group was not conditioned (injected with sodium chloride). To test for taste aversion conditioning, saccharin was offered for 20 min/day for 3 days. Nonconditioned BN and WKY rats consumed equal amounts of saccharin on test days. In both strains, conditioned rats showed a saccharin aversion. However, conditioning was less robust in BN than in WKY rats. WKY rats showed good LI of the conditioned taste aversion in that preexposed WKY rats consumed significantly more saccharin on test days than conditioned, nonpreexposed WKY rats. Preexposed BN rats did not consume significantly more saccharin on test days than conditioned, nonpreexposed BN rats. The previously reported deficiency in PPI in the BN rats was confirmed here 1 week after the taste aversion experiment. These results suggest that BN rats show deficient LI as well as PPI and display poor associative learning, a trait also reported in schizophrenia.

Spinal nicotinic receptor activity in a genetic model of hypertension.

Intrathecal cytisine, a nicotinic receptor agonist, elicits greater dose-dependent increases in blood pressure, heart rate and nociceptive responses in SHR than normotensive rat strains. Similar to adult rats, cardiovascular and nociceptive responses were augmented in prehypertensive SHR than age-matched WKY. While hydralazine or captopril pretreatment significantly lowered blood pressure in both SHR and WKY rats, responses to i.t. cytisine were still greater in SHR. By contrast, i.t. cytisine elicited responses were not exaggerated in DOCA-salt hypertensive WKY rats. Pressor and irritation responses to i.t. cytisine can be divided into a transient, initial and persisting, late phases. Both are augmented in SHR. In F1 rats, only the late phase pressor and pain responses to i.t. cytisine are similar in magnitude to those observed in SHR suggesting a possible dominant trait in the SHR. Overall, our findings suggest that hyper-responsiveness in nociception and pressor activity to spinal cytisine in SHR may be pathogenetically associated, but not a consequence, of hypertension.

Repeated restraint stress-induced increase in baroreceptor reflex sensitivity: role of corticotropin-releasing factor.

The effect of central administration of a corticotropin-releasing factor antagonist on repeated restraint stress-induced changes in the baroreceptor reflex response to phenylephrine was examined in male Wistar-Kyoto rats. Rats were instrumented with intracerebroventricular guide cannula and femoral arterial and venous catheters. On each of 5 consecutive days, two groups received either a central infusion of saline or an infusion of the corticotropin-releasing factor antagonist, astressin. One saline- and one astressin-treated group experienced 20 min of restraint stress 10 min after each infusion. The other saline- and astressin-treated groups served as non-stressed controls. Twenty-four hours later, each rat received 3 doses of phenylephrine which produced equivalent increases in mean arterial pressure in each of the 4 treatment groups. Reflex bradycardia was significantly greater in the saline-treated/repeated restraint group than in the saline-treated/no restraint group. This effect of repeated restraint on the baroreceptor reflex was attenuated by administration of astressin prior to each session of restraint. A single 20 min session of restraint stress failed to alter baroreceptor reflex sensitivity. However, repeated central infusions of exogenous CRF failed to alter BRR sensitivity. In a separate experiment, astressin failed to attenuate the increases in mean arterial pressure and heart rate which occurred during each session of restraint stress and, in fact, diminished habituation of the blood pressure response in the last session. The results suggest that repeated stress increases thesensitivity of the baroreceptor reflex and that corticotropin-releasing factor has a role in this stress-induced change.

Prepulse startle deficit in the Brown Norway rat: a potential genetic model.

Prepulse inhibition (PPI), an operational measure of sensorimotor gating, is deficient in schizophrenia patients. PPI was compared among 4 strains of rats: Sprague-Dawley, Spontaneously Hypertensive, Wistar Kyoto (WKY), and Brown Norway (BN). PPI was dramatically lower in BN versus the other strains, especially WKY, for both acoustic and airpuff startle stimuli, whereas startle amplitude was similar between BN and WKY. Female BN also had lower PPI than did female WKY. Response to increasing prepulse intensities showed a right shift in the BN relative to the WKY. Visual prepulses also showed deficiencies in BN versus WKY. The absence of background noise did not negate strain differences. Auditory brainstem response to clicks and tone pips revealed no differences in auditory threshold between the 2 strains. These results are the first to demonstrate that BN have impaired sensorimotor gating compared with WKY, without impaired acoustic acuity.

New target regions for human hypertension via comparative genomics.

Models of human disease have long been used to understand the basic pathophysiology of disease and to facilitate the discovery of new therapeutics. However, as long as models have been used there have been debates about the utility of these models and their ability to mimic clinical disease at the phenotypic level. The application of genetic studies to both humans and model systems allows for a new paradigm, whereby a novel comparative genomics strategy combined with phenotypic correlates can be used to bridge between clinical relevance and model utility. This study presents a comparative genomic map for "candidate hypertension loci in humans" based on translating QTLs between rat and human, predicting 26 chromosomal regions in the human genome that are very likely to harbor hypertension genes. The predictive power appears robust, as several of these regions have also been implicated in mouse, suggesting that these regions represent primary targets for the development of SNPs for linkage disequilibrium testing in humans and/or provide a means to select specific models for additional functional studies and the development of new therapeutics.

Airpuff startle stress elicited fos expression in brain cardiovascular areas of young SHR and WKY rats.

Prior studies comparing Fos expression in adult Wistar Kyoto (WKY) and Spontaneously Hypertensive rats (SHR) identified more Fos-positive neurons in a subset of brain regions following two stressors: placement in a startle chamber and presentation of an airpuff startle stimulus. The present study assessed Fos expression in five week old SHR and WKY rats in those same brain areas. Like adults, young SHR expressed more Fos-positive neurons than WKY in response to the startle chamber alone. Unlike adults, in the SHR only the locus coeruleus showed a increases in Fos expression following addition of the airpuff. Otherwise, startle chamber and airpuff startle treatments induced roughly equivalent Fos expression in the SHR, possibly reflecting a ceiling effect. Young WKY exhibited predominant airpuff-induced elevations. The present results demonstrate that certain brain regions are strain-differentially activated by stressors prior to overt hypertension and that differential Fos expression is an early developmental feature of these strains.

Strain differences in Fos expression following airpuff startle in Spontaneously Hypertensive and Wistar Kyoto rats.

The airpuff startle stimulus elicits both a behavioral and a concurrent sympathetic and parasympathetic activation, which have been shown to differ between inbred normotensive Wistar Kyoto and Spontaneously Hypertensive rat strains. Neither the brain sites responsible for the cardiovascular and motor responses, nor the origins of the strain differential responses, have yet been elucidated. The goals of the present study were (i) to define the neuronal pattern of immunoreactive Fos expression to the airpuff stimulus, and (ii) to determine whether this pattern of expression differed between the two contrasting inbred rat strains, thereby relating to observed differences in response. The airpuff stimulus induced Fos protein expression in discrete nuclei within the hypothalamus, thalamus, midbrain, pons and medulla of both strains, with strain-dependent differences evident in the hypothalamus (lateral, ventromedial and dorsomedial), pons (locus coeruleus) and medulla (rostroventrolateral medulla and solitary tract nuclei). To remove Fos expression arising from test chamber novelty, which was observed in both strains, a subset of animals was habituated to the test chamber for four days prior to testing. Habituation reduced Fos expression in several brain regions in the Wistar Kyoto, but failed to do so in the Spontaneously Hypertensive rat. The present results are the first to identify a set of brain regions likely to be responsible for the mediation of the cardiovascular and motor responses associated with the airpuff startle stimulus. Several of the identified areas contain neurotransmitters implicated by prior pharmacological studies. Further, these data identify differences in the degree of activation of specific neuronal structures that probably underlie strain differences in the cardiovascular response to the airpuff. Additionally, the results provide a cellular correlate to reported deficits in behavioral habituation by the Spontaneously Hypertensive rat and suggest a potentially profound difference between the ability of these two strains to adapt to repeated mild stress stimuli.

Catecholamine storage vesicle protein expression in genetic hypertension.

Chromogranin A expression is heritable in humans, and both plasma chromogranin A concentration and its releasable adrenal and sympathetic neuronal pools are augmented in established essential (hereditary) hypertension. To evaluate chromogranin A further as a simpler or "intermediate phenotype" in the complex trait of hypertension, we studied chromogranin A expression in the spontaneously hypertensive rat (SHR), a rodent model of essential hypertension. Both plasma (p < 0.0001) and adrenal medullary (p = 0.003 to p < 0.0001) chromogranin A were elevated in the SHR, even at the earliest stages (3-4 weeks of age). In the adult adrenal gland, both chromogranin A (p=0.005) and norepinephrine (p=0.011) were increased in the SHR, while dopamine beta-hydroxylase activity was diminished (p < 0.0001). Chromogranin A mRNA expression was also elevated in the SHR adrenal medulla (p = 0.017). Differences in chromogranin A processing were not noted between SHR and Wistar Kyoto control (WKY) rats. In an SHR x WKY genetic intercross, control of the adrenal chromogranin A phenotype by a single major locus was suggested by comparison of phenotypic variance of the F2 vs F1 generations, and by bimodal frequency histogram (3:1 ratio), confirmed by maximum likelihood analysis (chi2 = 74.6, p < 0.000001) in the F2 generation. However, microsatellite alleles at a surrogate locus (Ighe) 12.7 cM from chromogranin A (Chga), on rat chromosome 6, failed to co-segregate with blood pressure in an F2 generation (F = 0.06, p = 0.94). In another rodent model of hereditary hypertension, the genetically hypertensive mouse (BPH/2), adrenal chromogranin A (p=0.018) and norepinephrine (p = 0.004) were actually diminished. We conclude that over-expression of chromogranin A is a variable feature of mammalian genetic hypertension. In one rodent model (the SHR), over-expression of chromogranin A is largely controlled by a single genetic locus, but the chromogranin A locus itself is not directly linked to determination of the blood pressure elevation of the SHR.

Bradykinin-induced reductions in collagen gene expression involve prostacyclin.

Cardiac fibrosis after myocardial infarction and in chronic hypertension involves an increase in the synthesis and deposition of collagen within the myocardium. Angiotensin-converting enzyme (ACE) inhibitors limit hypertrophy and fibrosis; their mechanism of action remains controversial, although kinins have been implicated to play a role. Because both bradykinin and prostaglandins (PG) have been shown to reduce collagen gene expression in cardiac fibroblasts, the goal of this study was to determine whether the bradykinin effect was mediated through enhanced prostaglandin formation by cardiac fibroblasts. Bradykinin increased [3H]arachidonic acid metabolite release 2.3-fold over control and stimulated a dose-dependent increase in 6-keto PGF1alpha (the stable metabolite of PGI2) release from these cells, in which 1 nmol/L bradykinin produced a 4-fold increase in 6-keto PGF1alpha release. Beraprost (a PGI2 analogue) reduced steady-state proalpha1(I) and proalpha1(III) collagen mRNA levels by 35.6+/-6.6% and 34.2+/-10.0%, respectively. Bradykinin-induced reductions in collagen type I and III gene expression were reversed by pretreatment with indomethacin. Our results indicate that one mechanism by which bradykinin modulates collagen biosynthesis via the rabbit cardiac fibroblast involves formation of arachidonic acid metabolites, particularly PGI2. The results of the present study argue that stabilization of endogenous kinins (as by ACE inhibitors) would enhance prostacyclin production and result in the attenuation of collagen gene expression, with potential implications for collagen synthesis and deposition within the myocardium.

Attenuation of Fos expression to airpuff startle stimuli following tympanic membrane rupture.

The airpuff startle stimulus consists of two modalities, tactile and acoustic. Tympanic membrane rupture (TMR) effectively deafens a rat, thus preventing it from perceiving the acoustic component of the airpuff and permitting study of the tactile component in isolation. Previous studies have shown that the tactile modality is sufficient to drive the cardiovascular response to the airpuff, but cannot elicit the full behavioral startle response. In the present study Fos protein was used as a marker of neuronal activation to identify brain regions activated by the airpuff in both intact and TMR rats. Results show an attenuation of Fos expression following TMR in the dorsal and ventral cochlear nuclei, ventral nucleus of the lateral lemniscus and medial geniculate nucleus. In contrast, Fos expression following TMR was unchanged in the locus coeruleus, the laterodorsal tegmental nucleus, the supramammilary nucleus, and the ventromedial hypothalamic nucleus. Analysis of behavioral data confirmed that the startle response to the airpuff was diminished following TMR. These data are the first of which we know to employ an immediate early gene approach to discriminate between brain regions activated by the tactile and acoustic startle stimulus modalities. The results are discussed in terms of the classical acoustic startle circuit, and the central autonomic pathways activated by the tactile component of the airpuff.

Species variability in angiotensin receptor expression by cultured cardiac fibroblasts and the infarcted heart.

Cardiac fibroblasts, an abundant cell of the left ventricle (LV), proliferate and synthesize collagen in the heart after acute injury and during pressure overload hypertrophy. From many studies, angiotensin II (ANG II) receptors have been implicated in promoting collagen formation by the rat cardiac fibroblast. The present study examined species variability in ANG II receptor expression. Cultured rat fibroblasts expressed 43,000 +/- 15,000 ANG II (AT1-specific) receptors per cell (dissociation constant = 0.92 +/- 0.34 nM), whereas rabbit and neonate human cardiac fibroblast cultures expressed few receptors. Angiotensin increased intracellular Ca2+ concentration in rats but not in rabbit or human cardiac fibroblasts and stimulated arachidonic acid release in rat but not rabbit fibroblasts. In situ, 6 days after coronary artery ligation, angiotensin receptor expression was increased 34.8 +/- 13.4-fold in the infarcted area relative to the noninfarcted tissue in the rat LV, whereas rabbit hearts demonstrated only a 3.2 +/- 1.6-fold increase in ANG II binding within the infarcted tissue. These species differences in receptor expression raise questions as to the role of angiotensin as a mediator of collagen formation across species and as a direct target of angiotensin-converting enzyme inhibitors to regulate cardiac fibroblast function.

Prostacyclin release by rat cardiac fibroblasts: inhibition of collagen expression.

Cardiac fibroblasts, as the source of extracellular matrix for the left ventricle, subserve important functions to cardiac remodeling and fibrotic development following myocardial infarction or with pressure-overload cardiac hypertrophy. The fibroblast may be the target cell for angiotensin-converting enzyme inhibitors (ACEI) that are cardioprotective and reverse collagen deposition and remodeling but whose mechanisms of action remain controversial. Because we previously documented phenotypic differences between cardiac fibroblasts from the spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) left ventricle, the present study evaluated whether phenotypic differences also exist in the release of endogenous arachidonic acid metabolites or in the activation of phospholipase D, and the importance of observed differences to the formation of collagen and the mechanism of action of ACEI. The experimental design compared endogenous sources of arachidonic acid with exogenous prelabeling of cells. Angiotensin II stimulated greater arachidonic acid release than bradykinin, and WKY cells were more responsive than SHR. The major prostanoid formed by cardiac fibroblasts was prostaglandin I2 (PGI2), with more prostacyclin production by WKY cells than SHR cells both under nonstimulated conditions and in response to angiotensin II or bradykinin. Beraprost, a PGI2 analogue, was shown to decrease growth rate and DNA synthesis of fibroblasts and to inhibit mRNA expression for collagen types I and III, with SHR cells being less responsive to beraprost than WKY cells. These results potentially implicate eicosanoid metabolism, particularly PGI2, in collagen formation, fibrotic development, and cardiac remodeling, and they imply that the SHR genetic hypertension model may be predisposed to excess cardiac fibrosis.

Thiazide diuretics normalize urinary calcium in spontaneously hypertensive male rats.

The role of the thiazide-sensitive distal convoluted tubule (DCT) in the hypercalciuria of the spontaneously hypertensive rat (SHR) strain was examined by determining (a) the renal density of the thiazide diuretic receptor with 3H-metolazone, and (b) the renal response to a maximal dose of bendroflumethiazide (BFTZ). We confirm that the renal thiazide receptor density was greater in SHR than WKY (0.936 +/- 0.026 vs. 0.797 +/- 0.045 pmol/mg protein; P = 0.02). Prior to BFTZ the urinary excretion of calcium (0.525 +/- 0.061 vs. 0.274 +/- 0.049 micromol per micromol creatinine, P < 0.01) and sodium (12.6 +/- 1.27 vs. 7.89 +/- 0.926 micromol per micromol creatinine; P < 0.01) were greater in SHR versus WKY. BFTZ decreased the excretion of calcium only in SHR and to a level (0.250 +/- 0.032) not significantly different (P = 0.519) from WKY (0.225 +/- 0.032). Surprisingly, BFTZ increased chloride excretion to a greater extent in WKY than in SHR (P = 0.008). We postulate that hypercalciuria in SHR is a manifestation of incomplete uptake of calcium from the tubule lumen across the apical cell membrane in the DCT of the SHR nephron.

Locus coeruleus electrophysiological activity and responsivity to corticotropin-releasing factor in inbred hypertensive and normotensive rats.

The spontaneously hypertensive rat (SHR) and its normotensive progenitor, the Wistar-Kyoto rat (WKY), have been shown to be differentially responsive to the behavioral and endocrine effects of both stress and corticotropin-releasing factor (CRF), both of which increase locus coeruleus (LC) electrophysiological activity. However, the effect of central administration of CRF in these rat strains has yet to be examined. In the present studies, LC electrophysiological responsivity to intracerebroventricular infusions of CRF was assessed in SHR, an inbred strain of WKY rats (the WKY[LJ] rat), and an outbred normotensive rat strain, Sprague-Dawley (SD) rats. Spontaneous LC discharge rate, mean arterial blood pressure and heart rate were also examined. LC activity was increased to the same extent in the three rat strains in response to a 3 microg dose of CRF. However, WKY(LJ) rats showed an exaggerated LC in response to a 1 microg dose of CRF in comparison to the other rat strains tested at this dose. Spontaneous discharge rates of individual LC neurons were lower in both SHR and WKY[LJ] rats than in SD rats. Further, the variability of the discharge rates of LC neurons was greater in WKY[LJ] rats than in the other two strains. These results indicate that the WKY[LJ] rat may provide a useful model for assessing the role of sensitivity to CRF in stress responsiveness.

Angiotensinogen messenger RNA stabilization by angiotensin II.

OBJECTIVE: To further characterize the molecular mechanism whereby angiotensin II stabilizes the angiotensinogen messenger (m)RNA through binding studies of the previously isolated polysomal stabilizing protein to partial and mutagenized sequences of the 3' untranslated region of the gene and to explore its importance to rodent genetic hypertension. DESIGN: Analysis of angiotensinogen mRNA mutants for half-life and binding to a polysomal protein with a molecular weight of 12000. METHODS: Protein/RNA interactions were determined in band shift assays employing radiolabelled 3' untranslated region of angiotensinogen mRNA. Measurement of the mRNA half-life used a cell-free incubation system and 3' untranslated region DNA sequences were polymerase chain reaction (PCR) cloned and sequenced. Sequences of normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rat (SHR) strains were compared. Point mutations were introduced by site directed mutagenesis. RESULTS: The angiotensinogen 3' untranslated region exhibited specific binding to the polysomal 12000 molecular weight protein which, in an in vitro incubation system, increased 10-fold the half-life of full-length angiotensinogen mRNA; no effect was observed with 3' deleted angiotensinogen mRNA indicating a regulatory function of protein at the 3' untranslated region. Sequence analysis of PCR amplified DNA fragments identified a (G-->C) point mutation in the La Jolla colony SHR. Following introduction of this point mutation into wild-type 3' untranslated regions, protein binding significantly increased (wild-type binding constant, 19 mumol/l; mutant binding constant 3.5 mumol/l), indicating that this point mutation affects 3' untranslated region secondary structure, binding of the RNA stabilizing protein and, consequently, the half-life of angiotensinogen mRNA. Deletion of a U-rich region (position 1609-1613, UCCUU) expressed twice in the 3' untranslated region almost completely abolished protein binding suggesting this sequence as one part of the putative binding motif in the 3' untranslated region. CONCLUSIONS: Angiotensin II regulates hepatic angiotensinogen synthesis and secretion by inhibiting degradation of angiotensinogen mRNA by the action of a polysomal protein. Mutations in the 3' untranslated region mRNA coding sequence alter binding and half-life and may significantly affect the half-life of angiotensinogen mRNA thereby altering the secretion rate of angiotensinogen.

Spinal nicotinic receptor expression in spontaneously hypertensive rats.

Intrathecal administration of nicotinic agonists previously has been shown to result in exaggerated pressor and heart rate responses as well as greater nociceptive behavior in adult (12-week-old) spontaneously hypertensive rats (SHR) than in age-matched normotensive Wistar-Kyoto rats (WKY). Paradoxical to these augmented responses to nicotinic agonists in SHR, nicotinic receptor number in the spinal cord as measured by cytisine binding sites is lower in adult SHR than normotensive WKY and Sprague-Dawley rats. Using the high-affinity agonist epibatidine, we found similar differences in receptor number between strains in both in vitro ligand binding experiments with spinal cord membranes and in situ autoradiographic analyses. Spinal nicotinic receptor number did not differ in 5-week-old prehypertensive SHR and age-matched WKY; however, receptor numbers were higher in young rats of both strains than in their adult counterparts. Antihypertensive treatment (25 mg/kg per day hydralazine PO) in 6-week-old SHR from 6 to 12 weeks of age markedly reduced the progressive rise in blood pressure yet did not alter nicotinic receptor number compared with untreated rats. Similar treatment of WKY with hydralazine produced a slight fall in blood pressure but no change in receptor number. Thus, normalization of blood pressure by hydralazine in SHR does not result in a return of receptor expression to levels seen in normotensive rats. Higher centrally mediated pressor activity or augmented postcoupling events after neuronal nicotinic receptor stimulation may slowly downregulate expression of spinal nicotinic receptors in this genetically hypertensive rat strain.

Habituation of airpuff-elicited cardiovascular responses in the spontaneously hypertensive rat.

Repeated delivery of fast rise-time acoustic stimuli elicit cardiac changes in humans that reflect startle, orienting, and defense responses. To test the hypothesis that fast rise-time stimuli produce these responses in the rat, we evaluated magnitude, latency, and habituation of cardiovascular responses to brief airpuff stimuli in normotensive rats. We also evaluated airpuff-elicited cardiovascular responses in spontaneously hypertensive rats. In addition to a robust increase in blood pressure, airpuffs produced one or more of three sequential heart-rate responses in normotensive rats--first, short-latency tachycardia (latency ).8 s), then rapidly habituating bradycardia (latency 2.2 s), then long-latency tachycardia (latency 3.5 s)--which likely reflected startle, orienting, and defense responses, respectively. Airpuffs rarely produced bradycardia in hypertensive rats, suggesting that this strain does not appropriately orient to sensory stimuli. In addition, compared to normotensive rats, hypertensive rats exhibited greater between-session habituation of long-latency tachycardia and blood pressure increases. This finding contrasts with the Folkow hypothesis, which assumes that, in subjects with a genetic predisposition to develop hypertension, sympathetic responses will remain exaggerated after repeated stimulation, thus contributing to thickening of the arterial vasculature.

Central actions of corticotropin releasing factor on sensori-autonomic processing in normotensive and hypertensive rats.

The airpuff startle paradigm, a stress paradigm, elicits greater pressor and tachycardia responses in SHRLJ than WKYLJ which, unlike SHRLJ, also show bradycardia. CRF has been implicated in coordinating autonomic responses to stressors. The present studies explored the role of CRF, CRF antagonist D-PheCRF12-41 and chronic hydralazine in stress-induced cardiovascular responses. In SHRLJ CRF unmasked a bradycardia response, equal to WKYLJ receiving either aCSF or CRF, which was blocked by D-PheCRF12-41. CRF given to hydralazine-treated SHRLJ did not unmask a bradycardia. In WKYLJ CRF specifically lowered the pressor response on trial 1 compared to control, an effect abolished by the antagonist. WKYLJ receiving D-PheCRF12-41 prior to testing showed greater pressor and bradycardia responses when compared to control. In the WKYLJ CRF appears to exert an inhibitory role on the pressor and bradycardia responses while in the SHRLJ CRF exerts a permissive role on the bradycardia response which is lost when blood pressure is normalized with chronic hydralazine. CRF and D-PheCRF12-41 modify stress-induced cardiovascular responses in a strain-dependent dissimilar fashion. We conclude CRF has cardiovascular importance in the genetic hypertensive rat.

Intrathecal nicotinic agonist-elicited release of excitatory amino acids as measured by in vivo spinal microdialysis in rats.

The nicotinic agonists, nicotine and cytisine, when administered intrathecally elicit an increase in blood pressure and heart rate as well as a nociceptive response. By using a novel microdialysis procedure, in which a 4-cm dialysis tubing is inserted proximal to the intrathecal injection site, we have examined the release of excitatory amino acids associated with agonist stimulation. The nicotinic agonists, nicotine, cytisine and epibatidine, elicit dose-dependent increases in spinal release of Asp and Glu. The rank order of potencies of the nicotinic agonists in eliciting the cardiovascular and irritation responses correlates with the order of agonist potency in inducing Asp and Glu release in spinal microdialysates. In addition, a correlation is observed between the nociceptive and blood pressure response evoked by the nicotinic agonists and the spinal release of the excitatory amino acids. By examining the position of the permeable dialysis surface in relation to the agonist injection port, we found that the response to nicotine is localized to 5 to 10 mm distances, whereas the cytisine response may be elicited over longer distances. The marked desensitization observed upon repeated administration of cytisine is also reflected in diminished amino acid release. Amino acid release by nicotine can be antagonized by a channel blocking antagonist, mecamylamine, and by a competitive antagonist, dihydro-beta-erythroidine. Mecamylamine also inhibited the amino acid release elicited by epibatidine and cytisine. Hence, excitatory amino acid release is an appropriate response marker for the cellular events associated with nicotinic receptor stimulation in the spinal cord.

Role of the D1A dopamine receptor in the pathogenesis of genetic hypertension.

Since dopamine produced by the kidney is an intrarenal regulator of sodium transport, an abnormality of the dopaminergic system may be important in the pathogenesis of hypertension. In the spontaneously hypertensive rat (SHR), in spite of normal renal production of dopamine and receptor density, there is defective transduction of the D1 receptor signal in renal proximal tubules, resulting in decreased inhibition of sodium transport (Na+/H+ exchanger [NHE] and Na+/K+ATPase activity) by dopamine. To determine if impaired D1 receptor regulation of NHE in proximal tubules is related to hypertension, studies were performed in a F2 generation from female Wistar Kyoto (WKY) and male SHR crosses. A D1 agonist, SKF 81297, inhibited (37.6 +/- 4.7%) NHE activity in brush border membranes of normotensive F2s (systolic blood pressure < 140 mm Hg, n = 7) but not in hypertensive F2s (n = 21). Furthermore, a D1 agonist, SKF 38393, when infused into the renal artery, dose dependently increased sodium excretion in normotensive F2s (n = 3) without altering renal blood flow but was inactive in hypertensive F2s (n = 21). Since the major D1 receptor gene expressed in renal proximal tubules is the D1A subtype, we determined the importance of this gene in the control of blood pressure in mice lacking functional D1A receptors. Systolic blood pressure was greater in homozygous (n = 6) and heterozygous (n = 5) mice compared to normal sex matched litter mate controls (n = 12); moreover, the mice lacking one or both D1A alleles developed diastolic hypertension. The cosegregation with hypertension of an impaired D1 receptor regulation of renal sodium transport and the development of elevated systolic and diastolic pressure in mice lacking one or both D1A alleles suggest a causal relationship of the D1A receptor gene with hypertension.