Enzymes

An important part of chemical neuroanatomy was historically based on the distribution of various enzymes and enzymatic markers. In the mid-20th century, Koelle and Friedenwald developed a simple and sensitive histochemical method for revealing acetylcholinesterase (AChE), the degradative enzyme for acetylcholine. The application of AChE staining has consequently proven very useful in distinguishing brain areas (Fig. 1), although failure to colocalize AChE-positive neurons with catecholamines, the then darlings of the neuroscience community, dampened the interest of researchers in enzyme expression in the human. A comprehensive delineation of the rat brain by Paxinos and Watson was done largely on the basis of AChE reactivity with Nissl staining used as a second criterion. In the past 30 years, AChE histochemistry was successfully used for delineations of the brain in many mammalian species. Most important, AChE histochemistry works well on the fresh (unfixed) postmortem human brain, which allows this method to be successfully applied to the neuroanatomical delineation of the human brain. For example, in 1995 Paxinos and Huang mapped the entire human brain stem using AChE reactivity as a primary chemoarch-itectonic criterion. AChE staining was also used in pathological studies of the brains of patients with Alzheimer's disease and was recently employed as a relatively simple method for revealing the organization of the human hypothalamus (Fig. 1).

Another example in which enzyme distribution was instructive was the identification of the neuronal circuitry containing catecholamines (dopamine, nora-drenaline, or adrenaline) by the localization of the enzymes related to catecholamine synthesis. Using immunohistochemical techniques, researchers determined distributions of tyrosine hydroxylase (TH), aromatic amino acid decarboxylase, dopamine b-hydroxylase, and phenylethanolamine-N-methyl-transferase and were able to the delineate the relevant nuclei and subnuclei in the brain. In contrast, initial studies that aimed at directly localizing catecholamine molecules through chemical reaction with aldehydes were greatly limited by an inability to distinguish between the different catecholamines. The application of enzyme immunohistochemistry allowed the identification of 15 groups of catecholaminergic neurons in the mammalian brain. These cell groups were not confined to traditional cytoarchitectonic boundaries and consequently were termed A1-A16 (there is no A3 cell group), extending throughout the mammalian brain from the medulla to olfactory bulbs. In the human, as in the rat, the majority (A1, A2, and A4-A10) of catecholaminergic neuronal groups were found in the brain stem, where, for example, tyrosine hydroxylase immunostaining has been used to deline ate the intermediate reticular zone. Four prominent tyrosine hydroxylase-positive catecholaminergic cell groups (A11-A15) are located in the hypothalamus (Fig. 2) and one (A16) is located in the substantia innominata of the ventral forebrain. The latter cell group is thought to be homologous to the rat's catecholaminergic cell group in the olfactory bulb. Subsequent work has shown that cell groups such as the Al and Cl catecholaminergic neurons are critical for autonomic control in experimental animals and also that these cell groups are strikingly similar in rat and human. Many studies used multiple markers to confirm a high degree of conservation in the chemical identity of brain stem neurons in general between rat, monkey, and human.

Neuronal nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) is an enzyme that synthesizes the interneuronal messenger nitric oxide, hence the reason for the current intense interest in this protein. With respect to neuroanatomy, histochemis-try reveals a restricted distribution of this enzyme in the brain, a feature that allows accurate identification of exquisitely detailed subnuclear structures. For example, NADPH-d reactivity has revealed the otherwise ambiguous supraoculomotor cap and the dorso-lateral subdivision of the periaqueductal gray in the midbrain of the rat, rabbit, cat, monkey, and human. This finding allowed reliable identification of the interspecies homology for these microscopic subnuc-lear structures. Interestingly, the supraoculomotor cap has also been identified as being AChE positive.

NADPH-d reactivity has also been shown to reveal specific neuronal groups in the hypothalamus, thalamus, and medulla. It is a primary marker for the islands of Calleja in the ventral forebrain and reveals neuronal subpopulations in the human cortex. As an enzymatic neurochemical marker, NADPH-d histo-chemistry holds significant potential in current neu-roanatomy and has been selected for delineation in popular atlases of rat and monkey brains.

Breaking Bulimia

Breaking Bulimia

We have all been there: turning to the refrigerator if feeling lonely or bored or indulging in seconds or thirds if strained. But if you suffer from bulimia, the from time to time urge to overeat is more like an obsession.

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