chiro-and myo-Inositols are major components of the two inositol phosphoglycan mediators of insulin action. Previous work in this laboratory has shown hypo-chiroinositoluria in type H diabetic subjects and decreased chiroinositol in mediator prepared from skeletal-muscle biopsies of Pima Indian diabetic subjects together with increased myoinositol concentrations. Because mediator bioactivity was not previously examined, we decided to isolate the two types of insulin mediator from hemodialysate, urine, and autopsy muscle to investigate their bioactivity in control and type H diabetic subjects. Human mediator fractions were isolated at pH 2.0 and pH 1.3 from hemodialysate, urine, and autopsy muscle of type II diabetic subjects and nondiabetic control subjects. Mediators were assayed for bioactivity, and the relative chiroinositol/myo-inositol concentration ratio was determined for the mediator pH 2.0 samples by using HPLC or GC/MS. Regardless of source, the chiro-inositol-containing mediator pH 2.0 fractions from type H diabetic subjects were markedly less active than those from controls (50% or less) (P < 0.05).In addition, the chiro-inositol/myo-inositol ratio in samples from type H subjects was signficantly reduced (1/3-1/9) compared with controls (P < 0.05 for hemodialysate and P < 0.01 for muscle samples). In contrast, no difference in bioactivity was seen in myo-inositol-containing mediator pH 1.3 samples isolated from the same type II diabetic and control subjects. In type H diabetes there is a generalized deficiency of chiro-inositol mediator in the body in terms of both decreased chiro-inositol mediator (pH 2.0) bioactivity and chiro-inositol content.

Deproteinized skeletal muscle extracts free of major nucleotides from control and insulin-treated rats were fractionated and assayed for inhibition of protein phosphorylation by cyclic adenosine monophosphate (AMP)-dependent and -independent protein kinases. A differential effect of insulin on a particular fraction was observed on cyclic AMP-dependent protein kinase but not on cyclic AMP-independent protein kinases. This fraction that inhibited cyclic AMP-dependent protein kinase also stimulated glycogen synthase phosphoprotein phosphatase. It is proposed that this fraction may contain a mediator substance generateed in the presence of insulin.

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The spinal dorsal horn (SDH) and trigeminal subnucleus caudalis (Vc) contain second-order sensory neurons receiving primary afferent inputs from the somatic and orofacial regions, representing the first stage of nociceptive control in the central nervous system (CNS) and relaying nociceptive input to higher brain centers (Brown 1982;Dubner and Bennett 1983;Iggo et al. 1985;Sessle 2000). Vc is continuous with the cervical SDH and both SDH and Vc have a laminated structure (Brown 1982;Dubner and Bennett 1983;Iggo et al. 1985;Sessle 2000). Most small-diameter nociceptive primary afferent axons terminate in the superficial laminae of SDH and Vc (Dubner and Bennett 1983;Iggo et al. 1985;Sessle 2000 Here, we report that when compared with embryonic SDH neurons in culture, neurons isolated from the Vc region showed significantly slower growth, lower glutamate receptor activity, and more cells undergoing cell death. SDH neuron development was inhibited in co-cultures of SDH and Vc tissues while Vc neuron development was promoted by co-culture with SDH tissues. Furthermore, we identified that small (non-protein) ninhydrin-reacting molecules purified from either embryonic or post-natal Vc-conditioned medium inhibited neuronal growth whereas ninhydrin-reacting molecules from SDH-conditioned medium promoted neuronal growth. These findings suggest the involvement of locally released factors in the region-specific regulation of neuronal development in Vc and SDH, central nervous system regions playing critical roles in pain, and point to novel avenues for investigating central nervous system regionalization and for designing therapeutic approaches to manage neurodegenerative diseases and pain. Keywords: cell growth and death, hippocampus, local release, region-specific regulation, small (non-protein) ninhydrin-reacting molecules, spinal dorsal horn and medullary dorsal horn.