Role of the Adrenal Medulla in Hypoglycaemia-Associated Autonomic Failure-A Diabetic Perspective.

Hypoglycaemia-associated autonomic failure (HAAF) is characterised by an impairment in adrenal medullary and neurogenic symptom responses following episodes of recurrent hypoglycaemia. Here, we review the status quo of research related to the regulatory mechanisms of the adrenal medulla in its response to single and recurrent hypoglycaemia in both diabetic and non-diabetic subjects with particular focus given to catecholamine synthesis, enzymatic activity, and the impact of adrenal medullary peptides. Short-term post-transcriptional modifications, particularly phosphorylation at specific residues of tyrosine hydroxylase (TH), play a key role in the regulation of catecholamine synthesis. While the effects of recurrent hypoglycaemia on catecholamine synthetic enzymes remain inconsistent, long-term changes in TH protein expression suggest species-specific responses. Adrenomedullary peptides such as neuropeptide Y (NPY), galanin, and proenkephalin exhibit altered gene and protein expression in response to hypoglycaemia, suggesting a potential role in the modulation of catecholamine secretion. Of note is NPY, since its antagonism has been shown to prevent reductions in TH protein expression. This review highlights the need for further investigation into the molecular mechanisms involved in the adrenal medullary response to hypoglycaemia. Despite advancements in our understanding of HAAF in non-diabetic rodents, a reliable diabetic rodent model of HAAF remains a challenge.

The effects of recurrent hypoglycaemia and opioid antagonists on the adrenal catecholamine synthetic capacity in a rat model of HAAF.

In this study, we investigated the effects of recurrent hypoglycaemia on the adrenal catecholamine synthetic enzymes in a rat model of hypoglycaemia-associated autonomic failure (HAAF). We found that plasma adrenaline was significantly reduced by about 50% in response to recurrent hypoglycaemia versus single hypoglycaemia. However, tyrosine hydroxylase (TH) protein and phosphorylation at Ser31 and Ser40 were increased in HAAF; similarly, aromatic aminoacid decarboxylase protein was also increased indicating a likely increase in catecholamine synthesis in the adrenal gland. Opioid antagonists, naloxone and methylnaltrexone did not restore plasma adrenaline in HAAF; however, naloxone increased TH phosphorylation at Ser31 and Ser40.

Challenges in Modelling Hypoglycaemia-Associated Autonomic Failure: A Review of Human and Animal Studies.

Recurrent insulin-induced hypoglycaemia is a major limitation to insulin treatment in diabetes patients leading to a condition called hypoglycaemia-associated autonomic failure (HAAF). HAAF is characterised by reduced sympathoadrenal response to subsequent hypoglycaemia thereby predisposing the patients to severe hypoglycaemia that can lead to coma or even death. Despite several attempts being made, the mechanism of HAAF is yet to be clearly established. In order for the mechanism of HAAF to be elucidated, establishing a human/animal model of the phenomenon is the foremost requirement. Several research groups have attempted to reproduce the phenomenon in diabetic and nondiabetic humans and rodents and reported variable results. The success of the phenomenon is marked by a significant reduction in plasma adrenaline response to subsequent hypoglycaemic episode relative to that of the antecedent hypoglycaemic episode. A number of factors such as the insulin dosage, route of administration, fasting conditions, blood sampling methods and analyses, depth, duration, and number of antecedent hypoglycaemic episodes can impact the successful reproduction of the phenomenon and thus have to be carefully considered while developing the protocol. In this review, we have outlined the protocols followed by different research groups to reproduce the phenomenon in diabetic and nondiabetic humans and rodents including our own observations in rats and discussed the factors that have to be given careful consideration in reproducing the phenomenon successfully.

The Effects of Insulin-Induced Hypoglycaemia on Tyrosine Hydroxylase Phosphorylation in Rat Brain and Adrenal Gland.

In this study we investigated the effects of insulin-induced hypoglycaemia on tyrosine hydroxylase (TH) protein and TH phosphorylation in the adrenal gland, C1 cell group, locus coeruleus (LC) and midbrain dopaminergic cell groups that are thought to play a role in response to hypoglycaemia and compared the effects of different concentrations of insulin in rats. Insulin (1 and 10 U/kg) treatment caused similar reductions in blood glucose concentration (from 7.5-9 to 2-3 mmol/L); however, plasma adrenaline concentration was increased 20-30 fold in response to 10 U/kg insulin and only 14 fold following 1 U/kg. Time course studies (at 10 U/kg insulin) revealed that in the adrenal gland, Ser31 phosphorylation was increased between 30 and 90 min (4-5 fold), implying that TH was activated to increase catecholamine synthesis in adrenal medulla to replenish the stores. In the brain, Ser19 phosphorylation was limited to certain dopaminergic groups in the midbrain, while Ser31 phosphorylation was increased in most catecholaminergic regions at 60 min (1.3-2 fold), suggesting that Ser31 phosphorylation may be an important mechanism to maintain catecholamine synthesis in the brain. Comparing the effects of 1 and 10 U/kg insulin revealed that Ser31 phosphorylation was increased to similar extent in the adrenal gland and C1 cell group in response to both doses whereas Ser31 and Ser19 phosphorylation were only increased in response to 1 U/kg insulin in LC and in response to 10 U/kg insulin in most midbrain regions. Thus, the adrenal gland and some catecholaminergic brain regions become activated in response to insulin administration and brain catecholamines may be important for initiation of physiological defences against insulin-induced hypoglycaemia.

Investigation of tyrosine hydroxylase and BDNF in a low-dose rotenone model of Parkinson's disease.

Tyrosine hydroxylase (TH, the rate limiting-enzyme in catecholamine synthesis) is regulated acutely via phosphorylation of 3 serine residues--Ser19, 31 and 40, and chronically via changes in TH protein levels. In this study, we aimed to investigate how TH is regulated in the brain, gut and adrenal gland as well as changes in mature brain-derived neurotrophic factor (mBDNF) and proBDNF levels in a low-dose (2 mg/kg, 5 days/week for 4 weeks) rotenone model of Parkinson's disease (PD). Rearing behaviour decreased by week 3 in the rotenone group (p<0.01), with further decreases in rearing by week 4 (p<0.001); however, TH remained unchanged in the substantia nigra (SN) and striatum; TH levels were also unaltered in other catecholaminergic cell groups of the brainstem such as A1C1 neurons or locus coeruleus. In the olfactory bulb, TH protein decreased (2.5-fold, p<0.01) while Ser31 phosphorylation increased (1.4-fold, p<0.05) in the rotenone group. In contrast, TH protein was increased in the adrenal gland (2-fold, p<0.05) and colon (5-fold, p<0.05) of rotenone rats. mBDNF levels were not changed in the SN but were significantly reduced in plasma and significantly increased in the colon (2-fold, p<0.01) of rotenone-treated rats. This is the first study to assess TH and BDNF in the brain and periphery in the rotenone model before SN/striatum degeneration is evident. Together these results suggest that low-dose rotenone may have some potential to model the early stages of PD.