Emilio Badoer

Proceedings of the Australian Physiological and Pharmacological Society Symposium: The Hypothalamus HYPOTHALAMIC PARAVENTRICULAR NUCLEUS AND CARDIOVASCULAR REGULATION

1. The hypothalamic paraventricular nucleus (PVN) is an important integrative site within the brain composed of magnocellular and parvocellular neurons. It is known to influence sympathetic nerve activity.

Cardiac afferents play the dominant role in renal nerve inhibition elicited by volume expansion in the rabbit

In the rabbit, vagotomy combined with arterial baroreceptor denervation abolishes the renal sympathoinhibition elicited by volume expansion. However, the effect of removing cardiopulmonary afferents alone has not been investigated. The aim of the present study was to determine the role of the cardiac afferents in the renal sympathetic response elicited by volume expansion in the normal conscious rabbit. Four experimental groups were used in which rabbits were administered 1) volume expansion (Haemaccel, 1.9 ml/min for 60 min), 2) volume expansion + bolus intrapericardial procaine (20 mg) to block cardiac afferents, 3) volume expansion + intravenous procaine (20 mg bolus), and 4) intrapericardial procaine alone (20 mg bolus). Volume expansion did not significantly affect mean arterial pressure or heart rate but produced a profound fall in renal sympathetic nerve activity (∼50%). Intrapericardial procaine administered 30 min after the start of volume expansion markedly reversed the renal sympathoinhibition to within 20% of the pre-volume expansion level, an effect that wore off over 25 min. In contrast, intravenous procaine lowered renal sympathetic nerve activity slightly further. The results suggest that cardiac afferents play the dominant role in the renal sympathoinhibition in response to volume expansion in the normal conscious rabbit.In the rabbit, vagotomy combined with arterial baroreceptor denervation abolishes the renal sympathoinhibition elicited by volume expansion. However, the effect of removing cardiopulmonary afferents alone has not been investigated. The aim of the present study was to determine the role of the cardiac afferents in the renal sympathetic response elicited by volume expansion in the normal conscious rabbit. Four experimental groups were used in which rabbits were administered 1) volume expansion (Haemaccel, 1.9 ml/min for 60 min), 2) volume expansion + bolus intrapericardial procaine (20 mg) to block cardiac afferents, 3) volume expansion + intravenous procaine (20 mg bolus), and 4) intrapericardial procaine alone (20 mg bolus). Volume expansion did not significantly affect mean arterial pressure or heart rate but produced a profound fall in renal sympathetic nerve activity (approximately 50%). Intrapericardial procaine administered 30 min after the start of volume expansion markedly reversed the renal sympathoinhibition to within 20% of the pre-volume expansion level, an effect that wore off over 25 min. In contrast, intravenous procaine lowered renal sympathetic nerve activity slightly further. The results suggest that cardiac afferents play the dominant role in the renal sympathoinhibition in response to volume expansion in the normal conscious rabbit.

Efferent neural projections of angiotensin receptor (AT1) expressing neurones in the hypothalamic paraventricular nucleus of the rat.

Angiotensin II acts within the hypothalamic paraventricular nucleus (PVN) to help mediate a number of autonomic and endocrine responses. Evidence is sparse in regard to the particular neuronal cell groups that exhibit angiotensin II type 1 receptors within the PVN, and does not exist in relation to specified efferent neuronal populations in the nucleus. In the present experiments, retrogradely transported neuronal tracers were utilized in conjunction with immunohistochemistry using a well characterized polyclonal antibody raised against a decapeptide sequence at the carboxy terminus of the AT1 receptor, to determine whether it is preferentially distributed amongst different efferent populations within the PVN. The AT1 receptor is not associated with neurones in the PVN that project axons to the spinal cord, dorsomedial or ventrolateral medulla but coexists strongly with neurones in the anterior parvocellular division of the nucleus which direct axons to the median eminence. Such neurones often contain corticotropin releasing factor. These findings highlight the role that angiotensin II and AT1 receptors in the PVN may play in the mediation of responses to stress.

Microglia activation in the hypothalamic PVN following myocardial infarction

Following a myocardial infarction (MI), inflammatory cytokines are elevated in the brain, as well as in plasma, indicating that inflammation is occurring in the brain in addition to the periphery. Microglia are the immune cells in the central nervous system and can produce cytokines when they are activated by an insult or injury. In the present study, we investigated whether MI in rats induces activation of microglia in the brain. We used immunohistochemistry to detect CD11b (clone OX-42) and morphological changes to identify activated microglia. Compared to control rats that had undergone sham surgical procedures, there was a significant increase in activated microglia in the hypothalamic paraventricular nucleus (PVN) following myocardial infarction. Activated microglia were not observed in the ventral hypothalamus, adjacent to the PVN, nor in the cortex, indicating the response was not the result of a generalized inflammatory reaction in the brain. Echocardiography and haemodynamic parameters after myocardial infarction indicated that reduced left ventricular function but congestive heart failure had not developed. In conclusion, microglia are activated in the PVN but not in the adjacent hypothalamus following myocardial infarction. The activated microglia may contribute to the increased local production of pro-inflammatory cytokines observed in the PVN after myocardial infarction and resulting in reduced left ventricular function.

Effect of western and high fat diets on memory and cholinergic measures in the rat.

Recent evidence shows an association between obesity and cognitive decline. The present study aimed to determine whether a very high fat (60%) or western diet can affect working or spatial memory in rats and whether the diet-induced cognitive impairment is linked to the level of acetylcholine in the brain. Three groups of male Long Evans rats were fed either chow, western diet (21% fat, 0.15% cholesterol) or a high fat diet (60% fat) for 12 weeks (n=12 per group). Body weight, food intake and blood pressure were measured weekly. Behavioural testing, novel object recognition and Y-maze were carried out at 12 weeks. At the end of the study brain choline acetyltransferase and acetylcholinesterase levels were estimated. Results showed that consumption of a western diet for twelve weeks impaired a rats spatial memory (p<0.05), and increased body weight, calorie intake, blood pressure and triglyceride levels. Conversely our high fat diet also impaired spatial memory (p<0.05) but this effect was independent of the rats body weight or blood pressure. No significant changes in brain acetylcholine markers were observed. In conclusion, diets with higher fat content impaired hippocampal-dependant memory, even when hypertension and obesity are absent; however the mechanism is still unclear.

Cardiovascular role of the major noradrenergic cell groups in the rabbit: analysis based on 6-hydroxydopamine-induced transmitter release

We examined the role of the noradrenergic (NA) neurons of the A1, A2, A1 + A2, A5 and A6 + A7 regions on mean arterial pressure (MAP) and heart rate (HR), by comparing the acute responses of chronically lesioned and sham-operated rabbits to intracisternal 6-hydroxydopamine (6-OHDA, 600 micrograms/kg) which induces central release of transmitter. We studied rabbits (1) with intact arterial baroreceptors (non-denervated) and (2) after sino-aortic denervation (SAD). The acute transmitter release response consisted of an early fall in MAP (observed in SAD rabbits) and a late rise in MAP (observed in both non-denervated and SAD rabbits). Medullary lesions had no effect on either MAP component, but A5 and A6 + A7 lesions attenuated both pressor and depressor responses. Normally the transmitter release-induced MAP responses are modified by baroreceptor feedback. The 6-OHDA-induced HR changes were vagal in non-denervated rabbits and were sympathetically mediated in SAD rabbits. In non-denervated rabbits, A1, A2 and A1 + A2 lesions affected mainly the early vagal component, whilst A6 + A7 lesions affected the late vagal component. In SAD rabbits the early bradycardia was due to sympathetic inhibition and the late tachycardia due to sympathetic excitation; A1 + A2 lesions and A5 lesions attenuated the sympathetic bradycardia. We conclude that the various components of the MAP and HR responses are mediated through distinctive NA pathways; the deficits of a given lesion could be due to either to loss of NA cell bodies or of NA fibers of passage.

Role of the hypothalamic PVN in the regulation of renal sympathetic nerve activity and blood flow during hyperthermia and in heart failure

The hypothalamic paraventricular nucleus is a key integrative area in the brain involved in influencing sympathetic nerve activity and in the release of hormones or releasing factors that contribute to regulating body fluid homeostasis and endocrine function. The endocrine and hormonal regulatory function of the paraventricular nucleus is well studied, but the regulation of sympathetic nerve activity and blood flow by this region is less clear. Here we review the critical role of the paraventricular nucleus in regulating renal blood blow during hyperthermia and the evidence pointing to an important pathophysiological role of the paraventricular nucleus in the elevated renal sympathetic nerve activity that is a characteristic of heart failure.

nNOS-containing neurons in the hypothalamus and medulla project to the RVLM.

Nitric oxide (NO) within the brain is known to have an important influence on sympathetic nerve activity (SNA). NO is found in the paraventricular nucleus (PVN), caudal ventrolateral medulla (CVLM) and the nucleus tractus solitarius (NTS), regions that project to the rostral ventrolateral medulla (RVLM), an area that is critical in the regulation of SNA. The aim of the present study was to determine whether neurons in the PVN, NTS and CVLM that project to the RVLM contain the neuronal isoform of nitric oxide synthase (nNOS) and are, therefore, capable of producing NO. Under pentobarbitone general anaesthesia, the retrogradely-transported tracer, rhodamine-tagged microspheres, were microinjected into the RVLM of rats (n = 6). Two weeks later, the animals were re-anaesthetised, perfused with para-formaldehyde and the brains were removed. Hypothalamic and medullary sections were processed for nNOS immunohistochemistry and the RVLM-projecting neurons were identified using fluorescence microscopy. We found nNOS-containing neurons were present throughout the PVN, CVLM and NTS and that these were intermingled with neurons that projected to the RVLM. Of the neurons in the PVN and CVLM that projected to the RVLM, approximately 12 +/- 1% and 8 +/- 3%, respectively, contained nNOS. In the NTS only 1 +/- 1% of the neurons were double-labeled. This study highlights anatomical pathways emanating from the PVN and CVLM, in particular, which may contribute to the effects on SNA elicited by NO within the brain.

Microglia: Activation in acute and chronic inflammatory states and in response to cardiovascular dysfunction

Microglia are the resident immune cells in the central nervous system and are constantly monitoring their environment. After an insult, they are activated and secrete both pro- and anti-inflammatory mediators. Thus, they can have both detrimental and protective actions. Microglia are activated in many conditions that involve chronic inflammation such as Alzheimers and Parkinsons diseases and in neuropathic pain. Following cerebral ischemia and stroke, microglia are activated and acutely contribute to neuronal loss and infarct damage. Chronically, in this condition, neuroprotective actions of activated microglia include clearance of the dead cells and secretion of neurotrophins. Of great interest is the recent observation that following myocardial infarction, there is increased inflammation within the hypothalamus and a marked increase in activated microglia.

P2X purinoceptor subtypes on paraventricular nucleus neurones projecting to the rostral ventrolateral medulla in the rat

The rostral ventrolateral medulla (RVLM) is essential for the generation of sympathetic nerve activity. The RVLM receives a substantial innervation from the hypothalamic paraventricular nucleus (PVN). Activation of P2X purinoceptors via ATP has been shown to mediate fast excitatory synaptic neurotransmission. There is mounting evidence to suggest the presence of P2X purinoceptors in hypothalamic nuclei, including the PVN. In this study, we determined whether P2X1–P2X6 purinoceptor subtypes were present on PVN neurones that projected to the RVLM. Injection of the retrogradely transported tracer, rhodamine‐tagged microspheres, into the pressor region of the RVLM was used to identify the neurones in the PVN that innervated the RVLM. P2X1–P2X6 purinoceptors were detected by immunohistochemistry. Double‐labelled neurones were quantified and expressed as a proportion of the retrogradely labelled neurones. The proportions of double‐labelled neurones for each of the P2X purinoceptor subtypes varied, on average, from 14 to 29%. The P2X3 purinoceptor subtype was found to be the dominant purinoceptor subtype present on PVN neurones projecting to the RVLM. Additionally it was apparent that more than one P2X purinoceptor subtype was present on the PVN neurones projecting to the RVLM, since the sum of the average percentages of double‐labelled neurones for each P2X purinoceptor subtype exceeded 100%. These findings highlight the presence of the P2X1–P2X6 purinoceptors on PVN neurones projecting to the RVLM. The results suggest a potential role for ATP in the PVN in the regulation of sympathetic nerve activity.