This study was designed to assess the haem-peptide interactions which occur during progressive haemoglobin hydrolysis by digestive enzymes and their relationship with haem iron digestive absorption. The behaviour of different haemoglobin hydrolysates was studied using the Ussing chamber model. Hydrolysates were produced from enzyme digestion of bovine haemoglobin at pH 3 by pepsin and at pH 10 by subtilisin. Samples with increasing degrees of hydrolysis (0-15 %) were studied. Biochemical assays (pyridine haemochromogen method and UV absorption spectra) were used to follow haem solubility and haem-peptide interactions in samples. Increasing the hydrolysis level of haemoglobin was associated with an enhanced iron uptake; the highest uptake rate was reached between 8 and 11 % of globin hydrolysis, whichever enzyme was used. The mechanisms rendering iron soluble and available differ between the two enzymes. The comparison between biochemical and absorption data suggests that the formation of soluble peptide-haem complexes was not sufficient to enhance haem iron absorption, since globin-bound iron is poorly absorbed; an efficient absorption occurred only when haem was loosely bound to low molecular weight peptides.

Ionic mechanisms and signal transduction underlying noradrenaline (NA)-induced depolarization in single smooth muscle cells of guinea-pig vas deferens were studied. NA caused depolarization followed by action potentials through activation of α1-adrenoceptors. In the presence of nifedipine, no action potential was generated, and the magnitude of the depolarization depended on the concentration of NA (0.1-100 µm). NA, through α1-adrenoceptor activation, reduced the magnitude of membrane currents in response to voltage ramp pulses from -90 to -30 mV in a concentration-dependent manner. The reversal potential of the current inhibited by NA changed proportionally to the change in the equilibrium potential of K+, suggesting that NA inhibited K+ channel activity. Treatment of cells with GDPβS, an inhibitor of G proteins, or bisindolylmaleimide (BIM), a selective protein kinase C (PKC) inhibitor, prevented the NA inhibition of the currents. Application of 12-O-tetradecanoylphorbol 13-acetate (TPA), an activator of PKC, mimicked the effect of NA. It is suggested that in the smooth muscle of guinea-pig vas deferens, activation of α1-adrenoceptors and the subsequent activation of PKC led to inhibition of K+ currents, which is responsible for the depolarization induced by NA.

The purpose of the present study was to characterize the Ca2+ channels in smooth muscle cells from human stomach and to examine the effects of osmotic swelling on the channel activity. Ca2+ channel current with either Ca2+ or Ba2+ as charge carrier was recorded from freshly isolated smooth muscle cells using the conventional whole-cell patch clamp technique. The degree of cell swelling as a result of hypotonic challenge was monitored using a video image analysis system. The changes in intracellular Ca2+ concentration ([Ca2+]i) were measured by microfluorimetry. The pharmacological and voltage activation profile suggests a typical dihydropyridine-sensitive L-type Ca2+ current. Cell swelling, induced by hypotonic challenge, enhanced the amplitude of currents through L-type Ca2+ channels without significant effects on steady-state voltage dependency. After treatment with the L-type Ca2+ channel agonist Bay K 8644 (0.1-2 µM), no further significant increase in calcium channel current or corresponding [Ca2+]i transients were provoked by the swelling. The above results demonstrated that the presence of L-type Ca2+ current in smooth muscle cells of the human stomach and the augmentation of the current are closely associated with the volume increase resulting from hypotonic swelling. Experimental Physiology (2000) 85.5, 497-504.

The effects of osmotic stress on the cytosolic Ca2+ concentration ([Ca2+]i) in the endothelium of excised intact rat aorta were investigated using the indicator fura-2 and the patch clamp technique. Hyperosmotic stress evoked a reversible rise in endothelial [Ca2+]i in the presence but not absence of extracellular Ca2+, indicating that it evoked Ca2+ entry without release from intracellular stores. Hyposmotic stress was without significant effect. Cytochalasins B and D reduced the effect of hyperosmotic stress but not acetylcholine on endothelial [Ca2+]i. In endothelium isolated from underlying smooth muscle, hyperosmotic stress produced a rise in the [Ca2+]i and depolarisation of the endothelial membrane potential whereas hyposmotic stress was without effect. Mechanosensitive cationic channels recorded in cell-attached patches were activated by hyperosmotic solutions applied to the endothelium and inhibited by hyposmotic solutions. These data suggest that shrinkage of endothelial cells evokes an increase in [Ca2+]i by opening a pathway for Ca2+ entry from the extracellular space. The mechanosensitive ionic channels which we have previously described may be responsible for this response.

Isolated mammary arteries from ruminants were used in a conventional organ bath system. Acetylcholine relaxed bovine but not ovine mammary arteries; both types responded to sodium nitroprusside. Noradrenaline (NA) caused a dose-dependent increase in generated tension. An analogue of insulin-like growth factor I (long R3-IGF-I) caused a rightward shift in the NA response curve in bovine vessels with intact endothelium (P < 0.02), and also in sheep arteries (P < 0.01). In bovine vessels, this effect was abolished when the endothelium was removed. The effect of long R3-IGF-I in bovine vessels was abolished by N ω-nitro-L-arginine methyl ester (L-NAME) an inhibitor of nitric oxide synthase, suggesting the effect of IGF-I on mammary arteries in vitro requires NO generation. Experimental Physiology (2000) 85.3, 275-279.

Species differences in response to hypoxic damage have been observed in studies using whole hearts. The aims of this study were to determine whether (i) species differences in response to simulated hypoxia could be detected at the level of the single myocyte, and (ii) there were any interspecies differences in the Ca2+ handling properties of the cells. Ventricular myocytes were isolated from hearts of adult rats and guinea-pigs and electrically stimulated on the stage of a fluorescence microscope. Cell length was measured using an edge-tracking device, and total intracellular [Ca2+] ([Ca2+]i) determined using indo-1. Cells were exposed to metabolic inhibition (MI) (2.5 mM NaCN and no glucose) to simulate hypoxia followed by washout of CN and re-addition of glucose ('reperfusion'). Following exposure to MI, rat cells underwent rigor contracture in 18.8 ± 0.8 min (n = 80 cells), whereas the time was longer for guinea-pig cells (32.9 ± 1.2 min, n = 83) (P < 0.001). If cells were reperfused after 1-5 min in rigor, then rat cells showed improved morphological recovery compared with guinea-pig cells (P < 0.05); thereafter recovery decreased with increasing time spent in rigor, and was similar in both groups. In indo-1 loaded cells, [Ca2+]i was significantly increased in cells from both species at the end of MI; however, the actual increase was much higher in guinea-pig cells. Upon reperfusion, [Ca2+]i recovered fully in rat cells, but in guinea-pig cells there was no significant decrease. The restoration of [Ca2+]i to normal levels in rat cells following MI was associated with improved contractile recovery compared with guinea-pig cells. We conclude that rat cells are more resistant to effects of MI than are guinea-pig cells; this may be related to species differences in Ca2+ handling during and following exposure to MI. Experimental Physiology (2000) 85.5, 505-510.

The effects of cAMP accumulation evoked by acetylcholine (ACh) stimulation were studied in rat submandibular acinar cells by observing the exocytotic events, swelling of intercellular canaliculi (IC) and intracellular Ca2+ concentration ([Ca2+]i), which were monitored using an optical microscope. ACh stimulation evoked transient increases followed by sustained increases in the frequency of exocytotic events and IC swelling, while isoproterenol (isoprenaline; IPR) stimulation evoked sustained increases in these parameters. BAPTA treatment reduced the frequency of exocytotic events evoked by 5 µM ACh in the absence of extracellular Ca2+, and further addition of Rp-cAMPS or H-89 (protein kinase A (PKA) inhibitors) eliminated the remaining ACh-evoked responses (50 %). Addition of PKA inhibitors in the presence of extracellular Ca2+ reduced the frequency of exocytotic events evoked by 500 µM ACh in non-BAPTA-loaded cells. However, IC swelling evoked by 5 µM ACh was not affected by addition of PKA inhibitors, and was eliminated in BAPTA-loaded cells perfused with Ca2+-free solution. These results indicate that the IC swelling is regulated by [Ca2+]i and the frequency of exocytotic events is regulated by both [Ca2+]i and [cAMP]i during ACh stimulation. Addition of H-89 inhibited the capacitative Ca2+ entry into ACh-stimulated acinar cells. Biochemical analysis revealed that ACh stimulation increased the cAMP content in perfused submandibular glands. These results indicate that ACh stimulates the accumulation of cAMP in submandibular acinar cells and that this accumulation of cAMP modulates Ca2+-regulated exocytosis.

Shear stress is an important modulator of endothelial function which can affect vessel tone, endothelial morphology and gene transcription. However, the mechanism by which the endothelium detects changes in shear stress is the subject of conflicting reports. The possible contributions of changes in the cytosolic Ca2+ concentration ([Ca2+]i) and of membrane hyperpolarisation are disputed. Here we have investigated the effects of shear stress on these variables in native endothelium in situ in rat aorta and hepatic portal vein. Endothelial [Ca2+]i was recorded using the fluorescent indicator fura-2. The [Ca2+]i in unstimulated rat aortic endothelium was 86 ± 12 nM (n = 9). Acetylcholine (ACh) evoked relaxation of the aorta and a biphasic increase in endothelial [Ca2+]i. Shear stress that evoked relaxation comparable to that evoked by ACh did not affect the endothelial [Ca2+]i. Endothelial membrane potential was recorded using the patch clamp technique. The resting membrane potentials of rat aortic and portal vein endothelium were -47 ± 3.8 mV (n = 14) and -68 ± 4.5 mV (n = 9), respectively. Shear stress did not affect the endothelial membrane potential of rat aorta, but evoked a small (-1.6 ± 0.3 mV, n = 9) hyperpolarisation of the endothelium of rat portal vein in all preparations studied. These results suggest that neither changes in [Ca2+]i nor hyperpolarisation are involved in the reception of shear stress by native rat aortic endothelium in situ, but that shear stress induces small hyperpolarisations of endothelium of portal vein, a mechanism that may be present and of greater physiological significance in other parts of vascular system.

The most important tool for obtaining insight into the function of genes is the use of mutant model organisms. Homologous recombination in embryonic stem cells allows the systematic production of mouse mutants for any gene that has been cloned. Gene trap strategies have been designed to interrupt even unknown genes which are tagged by the inserted vector and can be characterised structurally and functionally. Complementary to such 'gene-driven' approaches, 'phenotype-driven' approaches are necessary to identify new genes or gene products through a search for mutants with specific defects, uncovering the function of genetic pathways in physiological and pathological processes. Mutagenesis using the alkylating agent N-ethyl-N-nitrosourea (ENU) is a powerful approach for the production of such mouse mutants. Since ENU induces mainly point mutations in premeiotic spermatogonia, this strategy allows the production of multiple alleles of a particular gene, which is pivotal for a fine tuned analysis of its function.

The relaxation effects of forskolin and methylxanthines on noradrenaline (NA)-induced contractions were investigated by measuring isotonic contraction and intracellular calcium concentration ([Ca2+]i) in the epididymal side of guinea-pig vas deferens. NA (100 µM) and high K+ (55 mM) induced a biphasic contraction; fast, transient (phasic) and slow, sustained (tonic) phases. Both phases in either NA or high K+ stimulation were abolished in Ca2+-free solution. Pretreatment with 10 µM nifedipine, an L-type Ca2+ channel blocker, reduced both phasic and tonic contractions induced by high K+. In the case of NA-induced contraction, however, nifedipine reduced the phasic contraction but not the tonic contraction. The nifedipine-insensitive tonic contraction was relaxed by the application of polyvalent cations (Mn2+, Co2+, Cd2+ and La3+). These findings indicate that NA-induced biphasic contraction is mainly due to nifedipine-insensitive Ca2+ influx, especially in the tonic phase. Cyclic AMP-increasing agents such as forskolin (0.5-10 µM), IBMX (5-500 µM) and caffeine (1-20 mM) relaxed the NA-induced contraction extensively in a concentration-dependent manner. However, these agents only partially relaxed the high K+-induced contraction. Forskolin (10 µM) and IBMX (100 µM) reduced the [Ca2+]i response to NA, but had no effect on the [Ca2+]i response to high K+. These results suggest that an increase in intracellular cAMP may relax the NA-induced contraction by attenuating a nifedipine-insensitive Ca2+ influx and by a mechanism independent of a reduction in [Ca2+]i.

The effect of chronic inflammation induced by Freund's Complete Adjuvant (FCA) on rat articular blood vessels and knee joint diameter was investigated. Blood flow changes in response to phenylephrine (an α1-adrenoceptor agonist) in FCA-treated and contralateral knee joints were studied over a 40 day period, using the laser Doppler flowmetery (LDF) technique. Unilateral injection of FCA (0.2 ml) increased the injected knee diameter on all days examined post-injection (P < 0.001) and its maximum increase (53 ± 2 %) was reached on day 3. After this, the diameter decreased gradually but did not return to its initial value. In control animals, topical application of 10-13-10- 7 mol phenylephrine onto the exposed joint capsule decreased blood flow dose dependently (11.1 ± 4.4 to 58.2 ± 4.5 %, respectively, P < 0.001). Unilateral injection with FCA attenuated the phenylephrine response in both ipsilateral and contralateral knees compared with the response of control animals (5.2 ± 1.6 to 48.3 ± 6.1 % and 1.9 ± 2.2 to 45.3 ± 5.6 %, respectively, P < 0.05). The reduction persisted for 3 weeks after FCA injection (ipsilateral for 21 days; contralateral for 30 days, P < 0.001). Subsequently the response returned towards normal. To avoid the influence of α2-adrenoceptors, yohimbine (an α2-adrenoceptor antagonist) was injected (0.5 mg kg-1, I.P.) 30 min before phenylephrine application. Yohimbine blocked the vasoconstrictor effect of 10-10-10-7 mol clonidine (an α2-adrenoceptor agonist, topical application) by 44-67.7 % inhibition, respectively (P < 0.001). Prazosin (an α1-adrenoceptor antagonist, 0.1 mg kg-1, I.P.) blocked the vasoconstrictor effect of phenylephrine (10-10-10-7 mol, topical application) effectively (42 to 69.8 % inhibition, respectively, P < 0.001). To assess the role of nitric oxide (NO) on the observed responses, N G-nitro-L-arginine methyl ester (L-NAME, NO synthase inhibitor) was applied topically (0.2 µmol) 5 min before phenylephrine application. L-NAME application at 7 and 14 days after FCA injection potentiated the vasoconstrictor response in the FCA-treated knee (P < 0.001) but had no significant effect on the contralateral knee. Blood pressure monitoring during phenylephrine, clonidine and L-NAME administration indicated that topical application of the drugs had no significant effect on the systemic blood pressure. These findings indicate that the vasoconstrictor response to phenylephrine was decreased in chronic inflammation and increased NO production could be involved.

Electrical stimulation of the parasympathetic auriculo-temporal nerve (40 Hz, 30 min), in the anaesthetized rat under α- and β-adrenoceptor blockade, increased [3H]thymidine and [3H]leucine uptake into the parotid glands by 80 and 263 %, respectively. The increase in response to parasympathetic stimulation was almost the same ([3H]thymidine 82 % and [3H]leucine 283 %) when atropine (2 mg kg-1 I.P. or I.V.) was included in the pretreatment. Neither intravenous infusion of vasoactive intestinal peptide (0.5-20 mg kg-1 min-1, 30 min) nor of bethanechol (10 mg kg-1 min-1, 30 min), under adrenoceptor blockade, increased the uptake of [3H]thymidine into the glands. However, these drugs increased [3H]leucine uptake, and in combination they interacted positively. Whereas vasoactive intestinal peptide is likely to be involved in the parasympathetic nerve-evoked protein synthesis, the nature of the non-adrenergic, non-cholinergic component(s) involved in the mitogenic response is presently unknown.

Embryonic stem (ES) cells are pluripotent cells isolated from the inner cell mass of blastocysts. ES cells are able to differentiate into the three primitive layers (endoderm, mesoderm and ectoderm) of the organism, including the germline. To study early stages of development, as well as to investigate the impact of a gene knock-out in vitro, ES cells are differentiated into three-dimensional structures called embryoid bodies, because of their ability to mimick post-implantation embryonic tissues. This review summarises the work on ES cell differentiation into haematopoietic and vascular cells, neuronal and glial cells, myocytes, and adipocytes, using this in vitro model of early embryogenesis. We also present the potential of this method to analyse the impact of genetic alterations in vitro

Secretion of fluid and proteins by salivary cells is under the control of parasympathetic and sympathetic autonomic nerves. In a recent study we have shown that, in the rat submandibular gland, autonomic nerves can also increase the secretion of IgA, a product of plasma cells secreted into saliva as SIgA (IgA bound to Secretory Component, the cleaved poly-immunoglobulin receptor). The present study aimed to determine if parotid secretion of SIgA is increased by autonomic nerves and to compare SIgA secretion with other parotid proteins stored and secreted by acinar and ductal cells. Assay of IgA in saliva evoked by parasympathetic nerve stimulation immediately following an extended rest period under anaesthesia indicated that it had been secreted into intraductal saliva in the absence of stimulation during the rest period. The mean rate of unstimulated IgA secretion (2.77 ± 0.28 µg min-1 g-1) and the 2.5-fold increase in IgA secretion evoked by parasympathetic stimulation were similar to results found previously in the rat submandibular gland. Sympathetic nerve stimulation increased SIgA secretion 2.7-fold, much less than in the submandibular gland. SDS-PAGE and Western blot analysis with anti-IgA and anti-Secretory Component antibodies confirmed that SIgA was the predominant form of IgA in saliva. Acinar-derived amylase and ductal-derived tissue kallikrein were more profoundly increased by parasympathetic and particularly sympathetic stimulation than SIgA. Overall, the results of the present study indicate that SIgA forms a prominent component of unstimulated parotid salivary protein secretion and that its secretion is similarly increased by stimulation of either autonomic nerve supply. The secretion of other parotid salivary proteins that are synthesized and stored by acinar or ductal cells is upregulated to a much greater extent by parasympathetic and particularly sympathetic stimulation. Experimental Physiology (2000) 85.5, 511-518.

Adenosine transport was characterized in human umbilical artery smooth muscle cells isolated from non-diabetic and diabetic pregnant subjects. Transport of adenosine was mediated by a Na+-independent transport system inhibited by nanomolar concentrations of nitrobenzylthioinosine (NBMPR) in both cell types. Diabetes increased adenosine transport, an effect that was associated with a higher maximal velocity (Vmax) for NBMPR-sensitive (es) saturable nucleoside transport (18 ± 2 vs. 61 ± 3 pmol (µg protein)-1 min-1, P < 0.05) and the maximal number of binding sites (Bmax) for specific [3H]NBMPR binding (74 ± 4 vs. 156 ± 10 pmol (µg protein)-1, P < 0.05), with no significant changes in the Michaelis-Menten (Km) and dissociation (Kd) constants, respectively. Adenosine transport was unaltered by inhibition of nitric oxide (NO) synthase (with 100 µM NG-nitro-L-arginine methyl ester, L-NAME) or protein synthesis (with 1 µM cycloheximide), but was increased by inhibition of adenylyl cyclase activity (with 100 µM, SQ-22536) in non-diabetic cells. Diabetes-induced adenosine transport was blocked by L-NAME and associated with an increase in L-[3H]citrulline formation from L-[3H]arginine and intracellular cGMP, but with a decrease in intracellular cAMP compared with non-diabetic cells. Expression of inducible NO synthase (iNOS) was unaltered by diabetes. Dibutyryl cGMP (dbcGMP) increased, but dibutyryl cAMP (dbcAMP) decreased, adenosine transport in non-diabetic cells. dbcGMP or the NO donor S-nitrosoacetylpenicillamine (SNAP, 100 µM) did not alter the diabetes-elevated adenosine transport. However, activation of adenylyl cyclase with forskolin (1 µM), directly or after incubation of cells with dbcAMP, inhibited adenosine transport in both cell types. Our findings provide the first evidence that adenosine transport in human umbilical artery smooth muscle cells is mediated by the NBMPR-sensitive transport system es, and that its activity is upregulated by gestational diabetes.

Salivary secretion of proteins from rat submandibular glands was studied using graded stimulation of the parasympathetic nerve in isolation, and then at a fixed rate in combination with graded sympathetic nerve stimulation. Increasing the frequency of parasympathetic nerve stimulation per se caused a gradual increase in the secretion of peroxidase (from acini) but only small increases in proteinase (from ductal cells) and IgA outputs. Dual stimulations, with an increasing frequency of sympathetic nerve stimulation on a background of low frequency parasympathetic nerve stimulation, showed that maximal acinar secretion of peroxidase required only a low frequency of additional sympathetic stimulation, whereas ductal secretion of kallikrein was greatest with the highest frequency of additional sympathetic stimulation (20 Hz in bursts). IgA secretion also required high frequency additional sympathetic stimulation in bursts for greatest output. Although a synergism occurred with parasympathetic plus sympathetic nerve stimulation for the secretion of both peroxidase and kallikrein it was not evident for the secretion of IgA. This presumably reflects a difference for exocytosis of proteins stored in granules (e.g. peroxidase and kallikrein) compared to those proteins continuously transported across the plasma membrane in vesicles by transcytosis. This work confirms that vesicular movement of secretory IgA can be increased by both parasympathetic and sympathetic nerve stimulation, but the frequency parameters differ for each nerve. Experimental Physiology (2000) 85.3, 281-286.

This study was undertaken to determine the reflex cardiovascular and respiratory responses to discrete stimulation of pulmonary arterial baroreceptors using a preparation in which secondary modulation of responses from other reflexes was prevented. Dogs were anaesthetised with α-chloralose, artificially ventilated, the chests widely opened and a cardiopulmonary bypass established. The main pulmonary arterial trunk, bifurcation and extrapulmonary arteries as far as the first lobar arteries on each side were vascularly isolated and perfused through the left pulmonary artery and drained via the right artery through a Starling resistance which controlled pulmonary arterial pressure. Pressures distending systemic baroreceptors and reflexogenic regions in the heart were controlled. Reflex vascular responses were assessed from changes in perfusion pressures to a vascularly isolated hind limb and to the remainder of the subdiaphragmatic systemic circulation, both of which were perfused at constant flows. Respiratory responses were assessed from recordings of efferent phrenic nerve activity. Increases in pulmonary arterial pressure consistently evoked increases in both perfusion pressures and in phrenic nerve activity. Both vascular and respiratory responses were obtained when pulmonary arterial pressure was increased to above about 30 mmHg. Responses increased at higher levels of pulmonary arterial pressures. In 13 dogs increases in pulmonary arterial pressure to 45 mmHg increased systemic perfusion pressure by 24 ± 7 mmHg (mean ± S.E.M.) from 162 ± 11 mmHg. Setting carotid sinus pressure at different levels did not influence the vascular response to changes in pulmonary arterial pressure. The presence of a negative intrathoracic pressure of -20 mmHg resulted in larger vascular responses being obtained at lower levels of pulmonary arterial pressure. This indicates that the reflex may be more effective in the intact closed-chest animal. These results demonstrate that stimulation of pulmonary arterial baroreceptors evokes a pressor reflex and augments respiratory drive. This reflex is likely to be elicited in circumstances where pulmonary arterial pressure increases and the negative excursions of intrathoracic pressure become greater. They are likely, therefore, to be involved in the cardio-respiratory response to exercise as well as in pathological states such as pulmonary hypertension or restrictive or obstructive lung disease.

The deterioration of the immune system with ageing, which leads to an increased morbidity and mortality from infections, appears to be related to decreases in specific lymphocyte functions. However, the alteration of non-specific immunity is a more controversial subject. Our purpose was to investigate the age-related changes of different functions of the non-specific immune response in peritoneal macrophages (adherence to tissues, mobility directed to a chemical gradient from an infectious focus or chemotaxis, phagocytosis of foreign agents and destruction of these agents by superoxide anion production) and in lymphocytes (adherence and chemotaxis) from peritoneum, axillary lymph nodes, spleen and thymus. We used young (12 weeks), adult (22 weeks), mature (48 weeks) and old (72 weeks) female BALB/c mice. The adherence capacity of macrophages and lymphocytes was greater in adult and old mice than in young animals. The chemotaxis of macrophages showed higher values in cells from young mice than in those from adult mice, increasing again in macrophages from mature and old animals. A similar behaviour was shown by phagocytosis, which reached its highest values in old animals. Anion superoxide production increased with age and again the highest values were obtained in the oldest mice. Conversely, chemotaxis of lymphocytes was higher in the adult and mature animals than in the young and old animals. We conclude that, although there is a decrease in lymphocyte chemotaxis in old animals, the non-specific immune response of macrophages instead of decreasing, may increase in aged mice with respect to the values seen in adult mice. Experimental Physiology (2000) 85.5, 519-525.

The present experiments were carried out in conscious lambs (1-2 weeks old, n = 9) and older sheep (11-12 weeks old, n = 11) to determine whether the cardiovascular and endocrine responses to 0, 10 and 20 % haemorrhage were developmentally regulated. The major novel finding of our study is that throughout the first 3 months of postnatal life, there is a similar decrease in mean arterial pressure and a similar restoration of pressure to pre-haemorrhage levels, for the same degree of blood loss, yet the mechanisms used to restore pressure appear to be age dependent as follows. In lambs, but not in older sheep, heart rate increased for 1 h after 20 % haemorrhage. Activation of the renin-angiotensin system was also greater and more prolonged in lambs than in older sheep following haemorrhage, and occurred at a lesser degree of blood loss. Plasma arginine vasopressin responses to haemorrhage were, however, similar in both age groups. These data provide new information that some of the mechanisms used to restore arterial pressure following blood volume depletion appear to be age dependent. Experimental Physiology (2000) 85.3, 287-293.

In the pathogenesis of autoimmune diseases, tolerance against self-determinants is lost and autoreactive lymphocytes are activated leading to pathological damage of single or multiple organs. Viral and other microbial infections have been implicated in these processes. Viruses may induce immunopathological damage by maintaining a chronic immune response against the locally persisting infectious agent. Alternatively, viruses may help to initiate anti-self immunoreactivity, e.g. by induction of an inflammatory milieu needed to overcome tolerance against self-antigens. Presentation of viral antigens and/or previously immunologically ignored self-antigens in secondary lymphoid organs is most probably the key event in the initiation of autoimmune diseases. Translocation of antigens to secondary lymphoid organs and primary induction of T cell responses is primarily mediated by dendritic cells (DCs). We discuss here two transgenic models of autoimmune diseases where DC-mediated antigen transport initiated autoimmune responses against microbial neoself antigens. In the first model, dose and timing of antigen delivery by DCs and turnover of antigenic peptides presented by DCs are the main parameters regulating the outcome of autoimmune diabetes. In the second model, chronic stimulation of organ-specific immune responses via DCs leads to severe cardiovascular immunopathology with arteritis, myocarditis and eventually dilated cardiomyopathy. Taken together, transgenic mouse models are valuable tools for delineating basic pathogenic mechanisms and evaluating therapeutic strategies to interfere with early detrimental processes that lead to manifest autoimmune diseases.