A LC-MS/MS method for the quantification of caffeine, betamethasone, clonidine and furosemide in cerebrospinal fluid of preterm infants.

BACKGROUND: Newborns, admitted to the Neonatal Intensive Care Unit (NICU), are exposed to a large number of medications, the majority of which are not labeled for use in infants, especially in preterm newborns, because clinical trials on their benefits and harms are lacking. There is a huge gap in knowledge on pharmacokinetic (PK) data in sick preterm infants, including that of drug penetration to cerebrospinal fluid (CSF). One of the issues is related to the lack of reliable analytical methods for the measurement of drugs in CSF. METHODS: In this paper we describe a specific and sensitive LC-MS/MS method for the simultaneous quantification in CSF of four commonly prescribed drugs in NICUs: caffeine, betamethasone, clonidine and furosemide. RESULTS: The method was validated following EMA guidelines and applied to several CSF samples of preterm infants with post-hemorrhagic ventricular dilatation in which a ventricular access device was applied. The range of the concentrations of the four drugs measured in the CSF was wide. CONCLUSIONS: Our method can be considered useful for further clinical studies to describe the PK aspects of these drugs in neonatal medicine.

Effects of Alzheimer's disease and normal aging on cerebrospinal fluid norepinephrine responses to yohimbine and clonidine.

BACKGROUND: The resting cerebrospinal fluid (CSF) norepinephrine concentration is unchanged or even increased in patients with Alzheimer's disease (AD). These in vivo findings appear to be inconsistent with the post-mortem locus ceruleus neuronal loss that is reported in patients with AD. METHODS: The effects of AD and advanced age on central nervous system noradrenergic status were estimated by comparing CSF norepinephrine concentrations following the administration of yohimbine hydrochloride, clonidine hydrochloride, and placebo in outpatients with AD and older and young normal subjects. Levels of yohimbine, its metabolite 11-hydroxy-yohimbine, and clonidine were measured in CSF and plasma samples. Behavioral responses were quantified by rating the Tension, Excitement, and Anxiety items on the Brief Psychiatric Rating Scale. RESULTS: Yohimbine-induced increases of CSF norepinephrine concentrations were greater in both patients with AD and normal older subjects than in normal young subjects. Clonidine-induced decreases of CSF norepinephrine concentrations did not differ among groups. Behavioral arousal following the administration of yohimbine was greater in patients with AD than in the other groups. CONCLUSIONS: Central nervous system noradrenergic responsiveness is enhanced in normal older subjects, and this age effect is retained in patients with AD. Behavioral sensitivity to increased central nervous system noradrenergic activity is enhanced in patients with AD.

Patterns of cerebrospinal fluid catechols support increased central noradrenergic responsiveness in aging and Alzheimer's disease.

BACKGROUND: High cerebrospinal fluid (CSF) norepinephrine (NE) concentrations in aging and Alzheimer's disease (AD) could reflect decreased NE clearance from central nervous system (CNS) extracellular fluid or increased release of NE into CNS extracellular fluid. Measuring CSF concentrations of the intraneuronal NE metabolite dihydroxyphenylglycol (DHPG), an estimate of NE clearance, and the NE precursor dihydroxyphenylacetic acid (DOPA), an estimate of NE biosynthesis, can help differentiate these mechanisms. METHODS: NE, DHPG, and DOPA were determined by HPLC in CSF and plasma obtained following yohimbine, clonidine, and placebo. Ten AD, 10 older, and 11 young subjects were studied. RESULTS: CSF DOPA following yohimbine was higher in older and AD than in young subjects. CSF DHPG did not differ among groups. Plasma DOPA following yohimbine was higher in AD than in young subjects. CONCLUSIONS: During alpha-2 adrenoreceptor blockade in both aging and AD, there are increased responses of CNS NE biosynthesis and release with unchanged CNS NE clearance. This pattern is consistent with partial loss of CNS noradrenergic neurons with compensatory activation of remaining CNS noradrenergic neurons. Given the marked loss of locus coeruleus (LC) noradrenergic neurons in AD, achievement of high CSF NE suggests particularly prominent compensatory activation of remaining LC neurons in this disorder.

Cerebrospinal fluid kinetics of epidural clonidine in man.

Ten patients with deafferentation pain after spinal cord injury were given 150 micrograms clonidine epidurally. CSF and plasma samples were collected over the following 24 h, and drug concentrations were measured by radio-immunoassay. The results of only 6 patients are included in the pharmacokinetic analysis because the catheters were not in the epidural space in the remaining 4 patients. These analyses revealed a mean maximum CSF concentration of 228 ng/ml whereas the mean plasma concentration at all time points was less than 0.7 ng/ml. The elimination half-life of epidural clonidine was 66 +/- 2 min, while the absorption half-life was 31 +/- 7 min, Tmax was 60 +/- 7 min and Cmax was 228 +/- 56 ng/ml. The ratio of the area under the curve (AUC) for CSF and plasma was 52. One patient's catheter was intrathecal and 3 were not in the epidural space. The measured plasma concentrations were similar after all injections. As 4 of 6 patients with epidural catheters obtained pain relief and all 3 patients with spasms obtained relief from epidural clonidine, these data suggest that clonidine has a place in the treatment of patients with spinal cord injury.

An antiserum to idazoxan recognizes an immunoreactive substance in human serum and cerebral spinal fluid which is not agmatine.

A polyclonal antibody was generated in rabbits to an idazoxan-albumin antigen. The anti-idazoxan antiserum had high affinity for unconjugated 3H-idazoxan (Kd of 19.8 nM) in a radio-immunoassay (RIA). Of various drugs and native molecules only idazoxan potently (Ki of 24 nM) inhibited 3H-idazoxan binding to the anti-idazoxan antibody. A few drugs weakly inhibited 3H-idazoxan binding (IC50 > 605 microM) with rank order of UK 14304 > guanabenz > cirazoline > amiloride > naphazoline. Neither agmatine, an endogenous clonidine displacing substance (CDS), catecholamines or imidazoles inhibited the binding of 3H-idazoxan to the anti-idazoxan antibody. The anti-idazoxan RIA was 4-6 fold more sensitive than an antibody to para-amino clonidine. The CDS detected by ligand displacement from bovine brain dose-dependently inhibited 3H-idazoxan binding. This immunoreactive (ir-) CDS activity was present in human (0.9-4.1 U/ml) and rat sera (1-2 U/ml) and in the cerebro-spinal fluid of eight patients with serious disease of the central nervous system, but not in controls. We conclude: (1) an anti-idazoxan RIA is a sensitive, selective and clinically applicable RIA for measuring ir-CDS; (2) ir-CDS is not agmatine; (3) CDS represents a family of endogenous ligands for imidazoline receptors including ir-CDS and agmatine.

Detection and measurement of an endogenous clonidine-displacing substance in human cerebrospinal fluid.

Clonidine-displacing substance (CDS) is a novel endogenous ligand for clonidine receptors previously detected in bovine brain and human serum. We examined for the first time whether CDS can be detected and measured in human cerebrospinal fluid (CSF). Using the [3H]clonidine displacement assay, we found that CDS could be identified and quantified in each of the CSF samples obtained from 81 patients with various neurological disorders. Mean level of CDS in CSF was 4.66 units/ml. Exceedingly high levels were observed in the CSF of patients with neoplastic meningitis (mean, 36.75 units/ml) and stroke (mean, 19.5 units/ml) (P < 0.0001). No correlation was found between CDS levels in CSF and age, gender, CSF protein or number of cells. CDS levels in CSF were higher than those in the serum (P < 0.01). We conclude that CDS is present and can be measured in human CSF. High CDS levels in CSF from patients with leptomeningeal metastases may serve as a tumor marker for malignant infiltration of the meninges. Additional studies in stroke patients will determine whether this endogenous ligand plays a role in the pathogenesis of cerebral ischemia.

Cerebrospinal fluid and plasma concentrations of clonidine in pigs after epidural, intravenous and intramuscular administration.

Epidural clonidine is an alternate way to treat severe pain in man. The cerebrospinal fluid (CSF) and plasma concentrations resulting after epidural, intravenous (i.v.) or intramuscular (i.m.) clonidine, 3 ug/kg b.w. have been determined by a sensitive gas chromatographic method. A porcine model was used, where the epidural and intrathecal spaces were cannulated via the atlanto-occipital membrane. After epidural administration of clonidine the CSF concentrations were maximal 20 minutes after the injection, 129 +/- 24 ng/ml (mean +/- S.D.). Clonidine was eliminated from CSF with an apparent half-life of 26 +/- 8.2 minutes (mean +/- S.D.). The plasma concentrations stayed below 1 ng/ml. Low plasma and undetectable CSF concentrations resulted after i.v. clonidine. Thus, epidural clonidine readily penetrates the dura mater into the CSF, and there is a marked gradient between CSF and plasma.

Computer-controlled epidural infusion to targeted cerebrospinal fluid concentrations in humans. Clonidine.

BACKGROUND: Pharmacokinetically designed infusions have been demonstrated to achieve rapidly and maintain desired concentrations of drug in plasma after intravenous administration. In this study we tested whether a similar approach, targeting concentrations in cerebrospinal fluid (CSF), could be used with epidural administration of the alpha 2-adrenergic analgesic clonidine. METHODS: After institutional review board approval and informed consent had been obtained, seven healthy volunteers received a clonidine infusion through a lower lumbar epidural catheter. Infusion of clonidine (10 micrograms/ml) was controlled by the STANPUMP program for sequential 75-min periods to targeted CSF clonidine concentrations of 25, 50, 75, and 150 ng/ml. Before reprogramming to the next higher targeted concentration, mean arterial blood pressure and heart rate were measured; blood was obtained for clonidine and catecholamine assays; and visual analog score for sedation and pain to immersion of foot and hand in ice water were obtained. CSF was collected during infusion with an indwelling lumbar intrathecal catheter and was analyzed for clonidine, catecholamines, and acetylcholine. RESULTS: CSF clonidine concentrations rapidly increased and were maintained at steady values with the stepped infusion, although observed concentrations were consistently greater than targeted. The relation between CSF clonidine concentration and analgesia in the foot was similar to that previously observed after epidural bolus administration. Clonidine also was associated with concentration-dependent sedation; decreased mean arterial blood pressure, heart rate, and CSF norepinephrine concentration; and increased CSF acetylcholine concentration. CONCLUSIONS: This study suggests that pharmacokinetically designed infusions of drugs in the epidural space in humans can maintain steady concentrations of drug in CSF. In addition to providing a useful tool for investigation of mechanisms of action and drug interactions, this technique may improve analgesia and diminish side effects from epidurally administered analgesics.

The efficacy of intrathecal morphine and clonidine in the treatment of pain after spinal cord injury.

We performed a double-blinded, randomized, controlled trial in 15 patients to determine the efficacy of intrathecal morphine or clonidine, alone or combined, in the treatment of neuropathic pain after spinal cord injury. The combination of morphine and clonidine produced significantly more pain relief than placebo 4 h after administration; either morphine or clonidine alone did not produce as much pain relief. In addition, lumbar and cervical cerebrospinal fluid (CSF) concentrations, sampled at these levels at different times after administration were examined for a relationship between pain relief and CSF drug concentration. Lumbar CSF drug concentrations were initially several orders of magnitude larger than those in cervical CSF. After 1-2 h, the concentrations of morphine in cervical CSF markedly exceeded those of clonidine. The concentration of morphine in the cervical CSF and the degree of pain relief correlated significantly. We conclude that intrathecal administration of a mixture of clonidine and morphine is more effective than either drug administered alone and is related to the CSF-borne drug concentration above the level of spinal cord injury. If there is pathology that may restrict CSF flow, consideration should be given to intrathecal administration above the level of spinal cord damage to provide an adequate drug concentration in this region.

Distribution, tolerability and tissue compatibility of intrathecal tizanidine in the sheep.

BACKGROUND: Tizanidine (TZD) is an alpha-2-adrenergic drug with potential spinal analgesic action and could be a substitute or additive for intrathecal (i.t.) opiates in chronic pain treatment. However, long-term tolerability and tissue compatibility are not yet established. METHODS: Three sheep were infused intrathecally with TZD up to 4000 micrograms/d over a time period of up to 440 d using implantable drug administration devices; one control animal was infused with vehicle only; standard values were collected from untreated sheep. CSF samples and blood samples were analyzed to determine pharmacokinetics and systemic redistribution. Behavioral effects were studied. The spinal cord tissue was investigated using standard laboratory histology. RESULTS: Bolus kinetics after i.t. injection of TZD are best described by a two-phase model. Elimination half-lives of TZD in CSF were 15 min and 152 min, respectively. Clearance of TZD from lumbar CSF was 0.48 ml/min. Doses higher than 500 microgram i.t. caused dose-dependent motor inactivity and sleepiness. Continuous i.t. TZD up to 4 micrograms/d was well tolerated regarding behavioral, physical activity, heart rate, muscle counts and total protein were detected both in saline and TZD-treated animals, presumably due to local irritation by the catheter and repeated sampling procedures. Histological evaluation of the spinal cord and adjacent tissues showed no abnormalities. CONCLUSION: The long-term intrathecal infusion of TZD is well tolerated and non-toxic in the sheep. The data favor clinical trials in patients with chronic pain.