Adrenal medulla and Parkinson's disease.

Parkinson's disease has been described as a multisystem disorder that includes alterations in the function of the autonomic nervous system. The activity of the adrenal medulla in this disease has not been thoroughly investigated. Previous reports are reviewed that demonstrate that the adrenal medullae of parkinsonian patients are compromised, having a decreased content of all catecholamines and several neuropeptides. An animal model was used to investigate whether the observations made in human patients were related to extended treatment with antiparkinsonian medications or were a natural concomitant of the disease. Administration of L-dopa and/or carbidopa to C57BL mice for 4-16 weeks had no significant effect on the level of any of the adrenal medullary catecholamines. Treatment with MPTP 4-16 weeks prior to sacrifice did not deplete adrenal medullary catecholamines in these animals, thus not fully mimicking Parkinson's disease in this animal model. The only significant effect was an interaction between group (MPTP or control) and treatment with antiparkinsonian medications; L-dopa, in the absence and presence of carbidopa, had opposite effects in the two groups. Based primarily on the lack of effect of antiparkinsonian medications on adrenal medullary catecholamines, it was concluded that the adrenal medullary depletion observed in human patients was a peripheral concomitant of Parkinson's disease.

The adrenal medulla and Parkinson's disease.

This paper reviews the literature describing the condition of the adrenal medulla in Parkinson's disease. Parkinson's disease is a neurodegenerative disorder that is characterized primarily by the loss of dopaminergic neurons in the substantia nigra. Clinical observations have revealed that Parkinson's disease is also frequently accompanied by a variety of autonomic symptoms. The adrenal medulla is a major component of the autonomic nervous system. However, until recently this organ has not been of particular interest in Parkinson's disease. Early studies found histologic abnormalities in adrenal medullary cells, and several groups measured urinary and plasma catecholamines to determine general autonomic status. In the late 1980s adrenal medullary tissue was first transplanted to the caudate nucleus in an attempt to augment the decreased levels of dopamine, and thus treat the symptoms of Parkinson's disease. At this time the status of the adrenal medulla in this disease became clinically important. We measured the total catecholamine content of the parkinsonian adrenal medulla in tissue collected both at autopsy and in conjunction with adrenal-caudate transplants. Adrenal medullary catecholamines and several neuropeptides were severely depressed in parkinsonian glands. Thus, the adrenal medulla appears to be a target of the peripheral manifestations of Parkinson's disease.

Increases in adrenolumbar venous flow and epinephrine secretion elicited by preoptic and hypothalamic stimulation in the cat.

Seventy-five sites were electrically stimulated within the preoptic and hypothalamic areas of anesthetized cats (n = 16). Adrenolumbar venous blood flow, secretion of epinephrine and norepinephrine, heart rate and mean arterial blood pressure were measured at baseline and following a standard stimulus (0.6 mA for 30 s) at each site. The most frequent adrenal medullary response to central stimulation was an increase in the secretion of both catecholamines (ng/min) greater than 10% of the baseline mean. Seventeen sites (23%) increased adrenolumbar venous flow greater than 12% above the mean baseline flow rate. This group of sites also evoked significantly greater increases in the secretion of epinephrine, and in mean arterial blood pressure and heart rate. These data support the hypothesis that secreted catecholamines, especially epinephrine, may act locally to increase blood flow through the adrenal medulla.

Adrenal medullary secretion with splanchnic stimulation in spinal cats.

This project was undertaken to determine whether previously observed adrenal medullary hyperactivity that developed following high spinal cord transection in the cat could be explained by increased sensitivity of the synapse between the splanchnic nerve and chromaffin cell. The splanchnic nerve was stimulated in acute (2-3 h; n = 7) or chronic (61-64 days; n = 7), spinally transected (T3) cats that were decerebrate and unanesthetized. Mean arterial blood pressure and adrenolumbar venous blood flow were significantly greater in the chronic animals. Stimulation (30 V; 1 ms pulses) was applied at 3 Hz and 30 Hz to deliver the same number of pulses within 3 min. Adrenal medullary secretion (ng/min) of epinephrine (EPI), norepinephrine (NE), dopamine, neuropeptide Y (NPY), [Met]enkephalin (ENK), and encrypted [Met]enkephalin was determined at baseline and in relation to both patterns of stimulation. With near threshold (3 Hz) stimulation, the following differences were observed between groups: (1) secretion of EPI, NPY, and ENK was significantly greater in the chronic than in the acute animals; and (2) preferential secretion of NE was elicited in the acute animals. These observations suggest that there may be some facilitation of the splanchnic nerve--chromaffin cell synapse that occurs over time following high thoracic spinal cord transection. However, it is likely that central, spinal mechanisms also contribute to adrenal medullary hyperactivity.

Decreased levels of [Met]enkephalin, neuropeptide Y, substance P, and vasoactive intestinal peptide in parkinsonian adrenal medulla.

Adrenal medullary tissue was collected from parkinsonian patients at autopsy and at the time of autologous transplantation of the adrenal medulla to the caudate nucleus, and from nonparkinsonian patients at autopsy and during nephrectomy. Levels of the following neuropeptides were measured by radioimmunoassay in samples of the medullary tissue: neuropeptide Y (NPY), substance P (SP), [Met]enkephalin ([Met]ENK), vasoactive intestinal peptide (VIP), peptide YY, and bombesin-like immunoreactivity. Regression analysis was used to establish a relationship between patient age, time to organ harvest, and peptide levels in nonparkinsonian tissue. Levels of [Met]ENK, VIP, NPY, and SP were significantly lower in parkinsonian adrenal medullae than that predicted from the control group. These results suggest that the adrenal medulla of a parkinsonian patient is severely compromised, either by the disease process itself or by the antiparkinsonian medications used to treat the symptoms of the disease.

Reduced D2 dopamine and muscarinic cholinergic receptor densities in caudate specimens from fluctuating parkinsonian patients.

Binding of spiperone and 3-quinuclidinyl benzilate (QNB), both labeled with hydrogen 3 (3H), were measured in caudate tissue obtained from 8 living parkinsonian patients at the time of cerebral transplantation. This was clinically homogeneous group of patients. All remained predominantly responsive to levodopa, although with marked disability secondary to clinical fluctuations (short-duration responses) and medication-induced dyskinesias; all were receiving substantial doses of levodopa and 6 of the 8 patients were additionally receiving bromocriptine or pergolide. Binding densities of dopamine D2 receptors, as measured by [3H]spiperone binding, were reduced in this group of patients, compared to caudate specimens from autopsy control subjects. This findings may reflect medication-induced receptor downregulation. Parallel changes occurred with muscarinic cholinergic receptors; [3H]QNB binding was significantly reduced, compared to autopsy control values. This reduction of muscarinic receptors might be due to loss of nigrostriatal terminals that are known to contain muscarinic receptors. Alternatively, muscarinic receptors may have been downregulated by increased corticostriatal glutamatergic input to cholinergic cells, inferred to be present based on the prominent levodopa-induced dyskinesias. Finally, receptor deficits could have been a reflection of more widespread degenerative cerebral disease, although levodopa-refractory symptoms were generally not pronounced in these patients.

Measurement of lumbar CSF levels of met-enkephalin, encrypted met-enkephalin, and neuropeptide Y in normal patients and in patients with Parkinson's disease before and after autologous transplantation of adrenal medulla into the caudate nucleus.

The levels in lumbar cerebrospinal fluid (CSF) of neuropeptide Y (NPY), methionine enkephalin (Enk), and Enk contained in amino- and carboxy-terminus extended forms (X-Enk) were examined in nine control patients undergoing elective surgical procedures and in eight patients with advanced Parkinson's disease, before and after the autologous transplantation of adrenal medullary fragments into the right caudate nucleus. The levels of CSF Enk and X-Enk before surgery in patients with Parkinson's disease were significantly less than those observed in control patients (Enk, 166 +/- 38 vs 264 +/- 44 pg/ml; X-Enk, 794 +/- 416 vs 1497 +/- 153 pg/ml). NPY levels did not differ (221 +/- 25 vs 193 +/- 23 pg/ml). After surgery, lumbar CSF samples were taken at 6 weeks, 12 weeks, 6 months, and 9 months. Placement of adrenal medullary fragments into the striatum had no effect on the levels of NPY or Enk at any time point. The levels of X-Enk were significantly enhanced only at 12 weeks (1138 +/- 140 pg/ml) but were at presurgical levels again by 6 months. These data suggest that the transplant was not functionally contributing to the CSF levels of these peptides.

Adrenal medullary transplantation into the brain for treatment of Parkinson's disease: clinical outcome and neurochemical studies.

Transplantation of adrenal medulla into the caudate nucleus as treatment for Parkinson's disease was performed in eight patients. Although our previous 6-month follow-up revealed early modest improvement, an extension of that follow-up to 1 year disclosed no additional gains in any patient. At the end of 1 year, only one patient could be categorized as moderately improved; three patients were mildly improved, and four patients were unimproved. The rationale for transplanting adrenal medulla was to reestablish a physiologic source of dopamine to the striatum. We measured cerebrospinal fluid (CSF) and plasma catecholamines and metabolites before and after transplantation. Conjugated dopamine (the predominant form of dopamine found in the CSF) and homovanillic acid (the major dopamine metabolite) were modestly and inconsistently increased in the CSF. Conjugated and free epinephrine and norepinephrine, as well as 3-methoxy-4-hydroxyphenylglycol concentrations were not increased in CSF after graft placement, an indication that the adrenal chromaffin cells were no longer producing high levels of these nondopamine catecholamines and metabolites. CSF cortisol concentrations were not increased after transplantation, compared with values from controls, consistent with low numbers of functioning adrenal cortical cells contaminating the graft (or poor survival). Posttransplantation CSF did not induce a neurotrophic effect in cell cultures of 15-day embryonic rat dorsal root ganglion or PC12 (rat pheochromocytoma) cell lines. Survival of samples of patients' adrenal medullary tissue for 2 weeks in tissue culture attested to the viability of the graft at the time of transplantation. The relative concentrations of dopamine to epinephrine or norepinephrine increased in these cultured adrenal medullary cells, presumably because of loss of the glucocorticoid influence on catecholamine synthesis. A wide variety of factors could have contributed to our failure to replicate the earlier impressive results of adrenal-to-brain transplantation reported by others. Continued transplantation studies in animal models of parkinsonism are necessary for better elucidation of these factors.

The secretion of catecholamines, chromogranin A and neuropeptide Y from the adrenal medulla of the cat via the adrenolumbar vein and thoracic duct: different anatomic routes based on size.

Secretion of the adrenal medulla was stimulated in nine cats by insulin-induced hypoglycemia. Levels of catecholamines (mol. wt 153-183), neuropeptide Y (mol. wt 4254) and chromogranin A (mol. wt 48,000) were measured in concurrently collected samples of adrenolumbar venous blood and thoracic duct lymph for up to 4 h following insulin administration. Insulin-induced hypoglycemia elicited an increase in the secretion of catecholamines, which reached peak levels in the adrenolumbar venous plasma at 1.5-2 h and in the lymph at 2.5 h. Although catecholamines were the most numerous measured molecules in the lymph, levels of norepinephrine and epinephrine were 75-250-fold less than those found in the adrenolumbar venous plasma. Neuropeptide Y in the adrenolumbar venous plasma reached peak levels between 1 and 1.5 h; at this time approximately 20% of the peak venous amount was detected in the lymph. Chromogranin A was found in approximately equal amounts in both plasma and lymph; the peak level in the plasma occurred at 1.5-2 h, while that in the lymph was reached at 2-3 h. We suggest that the size of a molecule influences the route it takes following exocytosis from the chromaffin vesicle. Smaller molecules such as catecholamines may pass directly into the circulation, while larger molecules such as chromogranin A may be temporarily sequestered in the interstitial space before passing into the lymph, and hence into the circulation.

Cerebrospinal fluid indices of blood-brain barrier permeability following adrenal-brain transplantation in patients with Parkinson's disease.

Cerebrospinal fluid (CSF) and serum or plasma concentrations of albumin, IgG and carbidopa were measured before and after adrenal-brain transplantation in patients with Parkinson's disease to indirectly assess blood-brain barrier (BBB) integrity. Previous studies in animals have suggested that the BBB is compromised by cerebral transplantation. CSF and plasma levodopa was also measured to permit comparison with the carbidopa values, recognizing that levodopa readily crosses the BBB via facilitated transport. Our patients underwent adrenal-brain transplantation in accordance with the method of Madrazo et al. (I. Madrazo, R. Drucker-Colin, V. Diaz, J. Martinez-Mata, C. Torres, and J. J. Becerril, 1987, N. Engl. J. Med. 316: 831-834) in which adrenal medullary pieces are implanted in the head of the caudate nucleus, in contact with the cerebrospinal fluid. All patients were maintained on oral carbidopa/levodopa therapy after surgery. CSF albumin/serum albumin and CSF IgG/serum IgG ratios were initially elevated above the preoperative baseline 6 weeks after the surgery; however, these values returned to the preoperative baseline by 6 months following the operation in six of seven patients. This suggested that the BBB was sufficiently intact to exclude these larger protein molecules from the CSF of these six patients. On the other hand, exogenously administered carbidopa, which normally is largely excluded from the cerebrospinal fluid by the BBB, was modestly increased in the CSF in four of the five patients in which it was measured. This suggests that the transplant BBB might be partially patent to small molecules for at least 6 months after the surgery. Whether increased passage of carbidopa into CSF and perhaps the transplant is of clinical significance has yet to be determined. Median CSF levodopa did not increase after surgery, probably because a limited defect in the BBB would be likely to be overshadowed by the effects of facilitated transport. CT scans performed following intravenous iothalamate meglumine contrast failed to reveal enhancement (dye leakage) near the transplantation site; however, artifact from the metal surgical clips used in the Madrazo procedure prevented good visualization of the area.

Catecholamines in CSF, plasma, and tissue after autologous transplantation of adrenal medulla to the brain in patients with Parkinson's disease.

Catecholamine concentrations were measured in tissue samples of caudate and adrenal medulla in eight patients with Parkinson's disease who were taking L-dopa and were undergoing autologous transplantation of adrenal medulla to caudate nucleus. High-performance liquid chromatography with electrochemical detection was used for the measurement of analytes. Dopamine concentrations were quite similar in the caudate and the adrenal medulla; epinephrine and norepinephrine concentrations were some 600 times and 90 times higher, respectively, than that of dopamine in adrenal medulla but were barely detectable in caudate nucleus. Catecholamines and metabolites were also measured, before and after transplantation, in lumbar cerebrospinal fluid (CSF) and plasma 1 hour after the patients' first morning dose of L-dopa. The major fractions of the catecholamines in CSF were sulfoconjugated. The concentrations of sulfoconjugated but not free dopamine were modestly increased in CSF after the transplantation, although plasma concentrations were unchanged. CSF concentrations of free and conjugated norepinephrine and epinephrine, 3-methoxy-4-hydroxyphenylglycol, and homovanillic acid were unchanged after the transplantation. The data suggest that the grafted tissue does not retain its noradrenergic or adrenergic properties after transplantation, and that dopamine formation in the brain may be modestly increased. Plasma catecholamines were unaffected after the removal of one adrenal gland for the transplant.

Decreased adrenal medullary catecholamines in adrenal transplanted parkinsonian patients compared to nephrectomy patients.

Adrenal medullary catecholamines were measured in tissue samples from eight patients who underwent autologous transplantation of the adrenal medulla to the caudate nucleus as a treatment for Parkinson's disease. These adrenal catecholamine levels were compared to a group of patients of similar age who underwent unilateral nephrectomy for renal cell carcinoma. The levels of each catecholamine, expressed as nanomoles per milligram wet weight tissue, were significantly lower (P less than or equal to 0.005) in the parkinsonian patients than in the nephrectomy patients. These observations support data reported previously from autopsy specimens and suggest that the adrenal medullae of parkinsonian patients may be a compromised source of dopamine-producing tissue; this may limit its effectiveness in eliciting maximum clinical improvement following transplantation.

Adrenal vein catecholamines and neuropeptides during splanchnic nerve stimulation in cats.

Splanchnic nerve stimulation in bursts at low (5 Hz) and high (50 Hz) frequency (30 V, 1 msec; train duration 1 sec; train rate 0.5/second) was employed in 10 cats under halothane anesthesia, during 10-minute periods, while blood samples were concurrently collected from the adrenal vein and femoral artery for the measurement of norepinephrine (NE), epinephrine (EPI), dopamine (DA), Met-enkephalin (ME), neuropeptide Y (NPY), peptide YY (PYY) and neurotensin (NT). In Group I (n = 5), splanchnic nerve stimulation was initially applied at 5 Hz followed after 20 min by a 50 Hz stimulus, while in Group II (n = 5) the stimulation sequence was reversed. Adrenal vein and femoral artery plasma levels of catecholamines and neuropeptides were not significantly affected by the stimulation sequence, while a significant decrease in blood pressure response was observed in Group II during the 5 Hz stimulation as compared to Group I, indicating desensitization. Splanchnic nerve stimulation at 5 Hz caused a preferential increase in adrenal vein NE (9-fold) versus EPI (7-fold) levels as compared to baseline, while 50 Hz stimulation led to further comparable increases in NE (5-fold) and EPI (6-fold) levels. Significant increases in adrenal vein DA and neuropeptide levels were only observed during 50 Hz stimulation, with DA showing a 5-fold, ME a 2.6-fold and NPY a 3-fold increase as compared to 5 Hz stimulation, and NT a 3.6-fold increase as compared to baseline. Present findings indicate different dynamics in the movement of catecholamines and neuropeptides from the adrenal.

Decreased catecholamine content in parkinsonian adrenal medullae.

Autopsy specimens of adrenal medullae from parkinsonian and nonparkinsonian patients were analyzed for free catecholamines by high-performance liquid chromatography with electrochemical detection. The total free catecholamine content (nanomoles free catecholamine per milligram protein) was significantly lower in the parkinsonian patients than in the control population when the values were corrected for age and time from death to organ harvest. It is not established whether this decreased catecholamine content in the adrenals of parkinsonian patients is a concomitant of the disease itself or whether it is secondary to drug therapies used to treat the symptoms of Parkinson's disease.

Adrenal medullary autograft transplantation into the striatum of patients with Parkinson's disease.

In eight patients with advanced Parkinson's disease, we performed autograft transplantation of adrenal medulla to the head of the caudate nucleus. Our technique was similar to that developed by Madrazo and co-workers in Mexico City. No major perioperative complications occurred except for somnolence in one patient for 8 days postoperatively. The follow-up period has been at least 6 months in seven of the patients, and only limited benefit has been apparent. The early morning Parkinson examination score in the "off" (unmedicated) state was significantly improved in one patient and slightly better in the other six. Diary card entries suggested a mild trend toward improvement (not statistically significant). Four of the seven patients were taking less levodopa 6 months after the operation than they had been preoperatively; three of five patients were no longer taking dopamine agonists postoperatively. We cannot exclude a placebo effect contributing to any of this improvement. A reduction in medication-induced dyskinesia was also noted, but this result may have been due to adjustments in doses or a slightly less potent effect of medication (or both factors). In summary, we have not yet been able to replicate the dramatic success reported for adrenal medullary transplantation by Madrazo's group, although our patients may have experienced mild to moderate improvement. We continue to maintain follow-up surveillance of these patients.

Effect of acute and chronic spinal transection on evoked secretion of adrenal medullary catecholamines in the cat.

Eight cats were spinally transected at T3. After an acute (0-5 days) or chronic (15-37 days) period, animals were rendered decerebrate and the effects of visceral (bladder distention) and somatic (sciatic nerve stimulation) stimuli were examined. Epinephrine, norepinephrine and dopamine levels were measured in plasma collected from the left adrenolumbar vein; heart rate and blood pressure were continuously monitored. In chronic animals both visceral and somatic stimuli most frequently evoked prominent increases in blood pressure and the secretion of adrenal medullary catecholamines; the same stimuli caused little change in these parameters in acute animals. These data indicate that a condition similar to the clinical syndrome of autonomic hyperreflexia may be elicited in the chronic spinally transected cat, and that this condition is accompanied by a notable activation of the adrenal medulla.

Increases in plasma catecholamines during naturally elicited defensive behavior in the cat.

Defensive behavior was evoked in 12 adult cats by the presentation of both interspecific (barking dog) and intraspecific (hypothalamically stimulated cat) stimuli. Each stimulus was presented 3 times. Concomitant activation of the sympatho-adrenal (SA) system was evaluated by radioenzymatic assay of plasma norepinephrine (NE) and epinephrine (E). The somatic and autonomic displays which accompanied defensive behavior were similar between stimuli, consisting of mydriasis, piloerection, growling, hissing and paw strikes. However, the sympatho-adrenal responses associated with interspecific and intraspecific defensive behaviors were different. Interspecific defensive behavior was accompanied by adrenal medullary release of both NE and E, which decreased with successive trials. Intraspecific defensive behavior was initially associated with release of E from the adrenal medulla; the pattern of catecholamine release in successive trials suggested that in later trials NE was released primarily from the sympathetic nerves. Furthermore, increases in plasma NE and E, as well as increases in heart rate and mean arterial blood pressure during the first trial were significantly greater when the behavioral stimulus was a dog. Although aspects of the feline display to threatening stimuli are stereotyped, the data suggest that the sympatho-adrenal response is more specific, and indicates preparation for fighting or escape.

Increases in blood flow from the adrenal gland following medial hypothalamic stimulation in the cat.

Medial hypothalamic sites previously shown to elicit either flight or defense were stimulated in the anesthetized cat. The rate of blood flow from the adrenolumbar (AL) vein ipsilateral to the side of stimulation was measured, as well as the adrenal output of norepinephrine and epinephrine (E). Stimulation at nearly 15% of the hypothalamic sites increased ipsilateral AL venous flow more than 20% over baseline rates. These sites also significantly increased heart rate, mean arterial pressure, and output of E from the ipsilateral adrenal gland. These observations suggest that hypothalamic sites where stimulation increased AL venous flow may overlap with regions that selectively mediate the release of E from the adrenal medulla.

Procedure for staining fixed human brain slices.

UNLABELLED: The described method provides a new technique for differentiating areas of gray and white matter in fixed human brain slices. The technique is a modification of an existing method permitting use of the nonfading copper phthalocyanine dye alcian blue. Stained slices show turquoise gray matter that contrasts sharply with areas of white matter. PROCEDURE: Cut human brains from gross anatomy laboratory cadavers into 4 mm slices and wash in running tap water for 12 hr. Oxidize slices in performic acid for 1.5 hr. Wash in running tap water for 12 hr. Stain slices in shallow dishes in 0.05% aqueous alcian blue. Wash in running tap water for 1 hr. Dry for 2-4 hr and embed in plastic.