Interactions between dopamine and glutamate play prominent roles in memory, addiction, and schizophrenia. Several lines of evidence have suggested that the ventral midbrain dopamine neurons that give rise to the major CNS dopaminergic projections may also be glutamatergic. To examine this possibility, we double immunostained ventral midbrain sections from rat and monkey for the dopamine-synthetic enzyme tyrosine hydroxylase and for glutamate; we found that most dopamine neurons immunostained for glutamate, both in rat and monkey. We then used postnatal cell culture to examine individual dopamine neurons. Again, most dopamine neurons immunostained for glutamate; they were also immunoreactive for phosphate-activated glutaminase, the major source of neurotransmitter glutamate. Inhibition of glutaminase reduced glutamate staining. In single-cell microculture, dopamine neurons gave rise to varicosities immunoreactive for both tyrosine hydroxylase and glutamate and others immunoreactive mainly for glutamate, which were found near the cell body. At the ultrastructural level, dopamine neurons formed occasional dopaminergic varicosities with symmetric synaptic specializations, but they more commonly formed nondopaminergic varicosities with asymmetric synaptic specializations. Stimulation of individual dopamine neurons evoked a fast glutamatergic autaptic EPSC that showed presynaptic inhibition caused by concomitant dopamine release. Thus, dopamine neurons may exert rapid synaptic actions via their glutamatergic synapses and slower modulatory actions via their dopaminergic synapses. Together with evidence for glutamate cotransmission in serotonergic raphe neurons and noradrenergic locus coeruleus neurons, the present results suggest that glutamatergic cotransmission may be the rule for central monoaminergic neurons.

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Frontal analysis continuous capillary electrophoresis was used to measure the binding of β-lactoglobulin (BLG) to sodium poly(styrenesulfonate) (PSS) and sodium poly(2-acrylamido-2-methylpropanesulfonate) (PAMPS), two strong polyanions with similar linear charge densities. The binding isotherms obtained were well-fit by the McGhee−von Hippel equation, yielding the intrinsic binding constant, K obs, and the binding site size, n, representing the number of polymer segments per bound protein. Two opposite ionic strength (I) dependencies of K obs for BLG−PSS were found depending upon pH, that is, increase of K obs with I at pH 7.0, and decrease of K obs with I at pH 6.3. The opposite I dependencies reflected the roles of electrostatic interactions for systems with heterogeneously charged components, but also demonstrated the inapplicability of a simple formulation (log K obs = log K° − Zφ log [M+]) put forward for the binding of protein to DNA. K obs for PAMPS was always much smaller than that for PSS at equal pH. In addition, n for BLG−PSS was small and independent of I and pH, while n for PAMPS was large and increased with I and pH, both results consistent with “tighter” binding of BLG to PSS than to PAMPS. This marked contrast may arise from the effects of polymer persistence length or from hydrophobic interactions.

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